Transfusion medicine changes of the paradigm

Transfusion medicine –changes of the paradigm

Prof. Primož Rožman, MD, PhD

Blood Transfusion Centre of Slovenia

Ljubljana, 2010

3

BTC

UniversityClinical Center

2500 beds7000 employees

300 employees

Dr. Janez Plečnik (1875 - 1940)

4

5

Contents – changes of the paradigms

EXPANSION OF TRANSFUSION MEDICINE 1990 - 20101

2

3 SAFETY: NAT, pathogen inact., leukodepletion, haemovigilance

4

CELLULAR THERAPIES, NEW PRODUCTS5

6 R&D PROJECTS & EXPERIENCES – BTC Ljubljana

MOLECULAR METHODS IN IMMUNOHAEMATOLOGY

ORGANISATION - LEGAL ASPECTS - TQM

Contents


2

3 SAFETY: NAT, pathogen inactivation, haemovigilance

4





8

CLASSICAL TRANSFUSION CHAIN

(VEIN – TO VEIN CONCEPT)

DONOR

BLOOD BANK

PATIENT

DONATION

PROCESSING

TESTING

APPLICATION

10

GENETICS:

HUGO 2001

TM DEVELOPEMENT - 3rd MILLENIUM

CLASSICAL TRANSFUSION

MEDICINE Stem cell

transplantation

Cellular therapies& tissue engineering

Gene therapy

Recombinant

technologies

METABOLOMIC

Viral /bacterial

inactivation

Blood

substitutes

GENOMICS/

TRANSCRIPTOMICS

PROTEOMICS

Protein microarrays TOTAL QUALITY

MANAGEMENT

MOLECULAR BIOLOGY

METHODS

Personalized medicine

Ex-vivo expansion

Haemovigilance

2020

2010

2000

1990

1980

Blood transfusion = cellular therapy !!

Blood bank = tissue establishment !!

Definition of transfusion medicine:

2010:

Contents


2

3 SAFETY: NAT, pathogen inactivation, haemovigilance

4





CAUSES FOR THE CHANGES OF BLOOD

SUPPLY ORGANISATION

14

• Globalisation

• Entering the EU – active partnership

European strategies – Lisboa declaration

• Laws, ethics – safety for every patient – EU

Directives on transfusion & cells/tissues

• New technologies

• Traffic and electronic communication

15

8,980

41,029

312

11,414

4,457

4,240

4,179

2,914

2,776

1,409

4,769

Trbovlje

Ptuj

Ljubljana

Blood transfusion services in Slovenia– 2010

H

H

H

•Collecting• X-matching

•collecting•processing•testing • X-matching

•collecting•processing•testing •NAT testing• X-matching• X-matching

Slovenian Red Cross

Blood donation in Slovenia is non-paid, voluntary and anonymous.

The Slovenian Red Cross organizes blood donation sessions with 56 local

organizations in the country.

Year 2004:

Slovenian Red Cross organized

1 070 blood donation sessions

323 on field collection sites

86 479 donations

First-time blood donors: 10 %

The Main Organizer of Blood Donation Sessions

Strategija ZTM Ljubljana

okt. 2008 – jan. 2010

19

20

21

22

KEY ELEMENTS FOR UNIFIED BLOOD

TRANSFUSION SERVICE IN SLOVENIA

• National blood supply policy

• National legislation – competent authority

• Information technology – national information

system • unified donor data base

• unified patient data base

• Telemedicine

23

24

Aims / expected results of telemedicine

• Equality of expertize nationwide

• Standardisation of operations

• Quality of service and blood supply regimens for

the whole state

• Improvement of:

Sampling results, archives

Connection to the laboratory robots

Transparency of data bases

Technical approach

+ teleconference

• High resolution

• Simple use

• Reproducible results

image capture

Graphic interface 3

Brežice

Before

Brežice

After: teleconsultation network 2010

Contents


2

3 SAFETY: NAT, pathogen inactivation, leukodepletion, haemovigilance

4





DIAGNOSTIC WINDOW

21

HIV

11 59 …

HBV

34 82 …

HCV

23

0 25 50 75 100 DAYS

AFTER INFECTION

DIAGNOSTIČ

WINDOW DETECTION OF ANTIBODIES ANTI-HIV - EIA

10 PCR

DIAGNOSTIC WINDOW DETECTION OF HBsAg - EIA

25 PCR

DIAGNOSTIC WINDOW DET. ANTI-HCV - EIA

59 PCR

NAT – nucleic acid testing in Slovenia

Since 2000 – HCV RNA PCR mini pools 48, Roche

COBAS Amplicor

Since 2007 – HCV, HBV, HIV – TMA method

(transcription mediated amplification), Chiron,

individual donor testing

Results:

2 HCV (window),

15 HBV (old occult HB infection),

0 HIV positive (in 100.000 donors per year)

32

PATHOGEN REDUCTION / INACTIVATION

1. Solvent-detergent treatment

2. Chemicals + UV light: preventing replication/

transcription of nucleic acids, inactivation of bacteria

and viruses

methylene blue

riboflavin (vitamin B2) - Mirasol (CaridianBCT)

psoralen-based chemicals - Intercept (Baxter/Cerus)

PATHOGEN REDUCTION/ INACTIVATION

Advantages:

Improved safety

Avoidance of bacterial detection

Prolonged storage

Precaution for emerging pathogens

Consistency with plasma

inactivation

Avoidance of irradiation

Disadvantages:

Loss of platelet count

Decrease of haemostatic platelet

function

No method is equally effective for

all organisms

Methods may be ineffective for

spore-forming organisms

Council of Europe’s opinion on platelet PR

PR is positive – should not be blocked

clinical transfusion trials for PR products needed

implementation of PR technologies should be

considered country by country, in relation to the

risks of transfusion

35

Pathogen reduction in Slovenia

2007 Decision – psoralen activation - InterceptBTC Medical board approvalDiscussion with cilinicians

2008 Buffy coat platelets productionAditive solution usagePremises adaptationDevices instalationStaff Training Intercept validationHV trainingRegulatory approval Domanović 2010

2009Clinical use

HV data collection

Pilot study of clinical effectivity

Production process

UNIVERSAL PRE-STORAGE LEUKODEPLETION

Improves preservation & storage

Reduced risks of:

febrile non-hemolytic reactions

transfusion associated infectious

diseases,

HLA sensitization, immunmodulation,

platelet refractoriness, transfusion

related GVHD

In Slovenia: from 2008 > all units are LD

39

HEMOVIGILANCE

40

Report of adverse reactions

to blood transfusion

Hemovigilance data -SLO

Adverse

reaction2003 2004 2005 2006 2007 2008 2009

Haemolysis 2 2 5 3 3 3 0

GVHD 0 0 0 0 0 0 0

TRALI/pulm.

Oed.

0 0 /2 0/3 1/12 0/14 1/11 2

PTP 0 0 0 0 0 0 15

Alergy/anap

hylaxis

41/1 54/6 55/4 67/2 68/3 82/5 70/3

NHTR 53 68 66 92 89 91 75

Bact/vir 0 0 4 1/1 0/3 0 1/1

Hypotension 1 3

Dyspnea 2 1

Other 9 14 12 12 12 8 3

Together 107 146 149 191 192 204 174

International Haemovigilance Network

42

• Australia* Austria* Belgium* Canada* Croatia* Denmark* Finland* France* Germany* Greece* Iceland* Italy* Japan•* Ireland

* Luxembourg* Malta* Norway* Portugal* New Zealand* Slovenia* Singapore* Sweden* South Africa* Spain* Switzerland* The Netherlands* United Kingdom* USA

- favour exchange of valid information between the members of the Network

- increase rapid alert / early warning between the members of the Network

- encourage joint activities between the members of the Network

- undertake educational activities in relation to haemovigilance.

Objectives

Contents


2

3 SAFETY: NAT, pathogen inact., leukodepletion, hemovigilance

4





Chromosomal localization

Rh1p34-36

H

9q34

HLA

GPIIa10p11.2

GPIV7q11

GPIa5q23.3

GPIc'

GPIb17p12

13 14 15 16 17 18 19 20 21 22 Y X

1 2 3 4 5 6 7 8 9 10 11 12

GPIIIa17q21-32

FcγRIIIb1q22

HNA AB0

HPA

Duffy1q22-q23

Kell7q33

Kidd18q11-q12

Molecular methods

• PCR-RFLP

• PCR-SSP

• PCR-SSO

• hybridization

• real-time PCR – TaqMan

• sequencing

• microarrays

• high troughput screening

RED CELLS

typing recently transfused or multitransfused patients

typing of patients with DAT+

resolving serological problems (weak D, partial D, acq.

phenotypes)

detection of weakly expressed Ag (Fyb-Fyx )

to distinguish allo- from autoantibody

46

PLATELETS

• refractoriness to platelet transfusion

• neonatal alloimmune thrombocytopenia (NAIT)

• post-transfusion thrombocytopenia HLA/HPA

•neonatal alloimmune neutropenia (NAN)

• transfusion related acute lung injury (TRALI)

GRANULOCYTES

Multitransfused patient – 1a

Original blood group: B; D-C+c+ E-e+; K-k+; Jk(a-b+); Fy(a+b+)02/B; Rh(d)/Rh(d); RhCe/Rhce; k/k; Jkb/Jkb; Fya/Fyb;

M/N; S/s

before transfusion

after transfusion

Multitransfused patients – 1b

Original blood group: B; D-C+c+ E-e+; K-k+; Jk(a-b+); Fy(a+b+)02/B; Rh(d)/Rh(d); RhCe/Rhce; k/k; Jkb/Jkb;

Fya/Fyb; M/N; S/s

- after 2. tr. event received 22 blood units- nonidentical units:

- 20x D+- 10x C-- 4x c-- 10x E+- 1x e-- 3x K+- 17x Jk(a+)- 7x Jk(b-)- 8x Fy(a-)- 5x Fy(b-)

before transfusion

after transfusion

control product

434 bp

specific product

bp

Contents


2

3 SAFETY: NAT, pathogen inactivation

4


6

7

R&D PROJECTS & EXPERIENCES – BTC Ljubljana



CELLULAR THERAPY

= transplantation of cells to replace/repair

damaged tissue and/or cells

51

Sources of cells:

• Hematopoietic progenitor cells (HPCs)

• Tissue stem cells.

• Embryonic stem cells/ iPSCs

• Ex vivo manipulated cells (DCs, CTLs, NKs, etc)

53

SOURCES OF THE STEM CELLS

Embrio

Odrasli

Fetus

Placenta

57

CLINICAL USE OF SCs

1. Haematological, genetic, malignant diseases

BM transplantation 1958

HSC from peripheral blood 1980ies

HSCs from UCB 1989

2. Regenerative medicine

Heart

Liver

CNS

Bone, cartilage, etc. . . . .

58

TYPES OF CELLS THAT CAN BE

USED FOR CELLULAR PURPOSES

IN BLOOD TRANSFUSION

1. Haematopoietic stem cells

2. Tissue stem cells

3. Embryonic stem cells

4. Ex vivo manipulated cells

5. In vitro cultivated cells

6. New generation SCs: iPS, transdiff.Cs

59

SOURCES:•Bone marrow

•Umbilical cord blood,

•Peripheral blood stem cell transplants

DIAGNOSES: •genetic disorders,

•leukemia and

•certain forms of cancer,

•CV diseases (CVI, MI, myocardiopathy, etc.)

• diseases of other organs/tissues

HAEMATOPOIETIC STEM CELLS

TISSUE STEM CELLS

60

SOURCES:

•Mesenchymal stem cells (BM, adipose tissue)

DIAGNOSES:

• rebuilding damaged cartilage in joints,

•repairing spinal cord injuries,

•autoimmune diseases (Mb Crohn, MS, …)

•neurological disorders

• several clinical trials going on

EMBRYONIC STEM CELLS/ iPSCS

61

SOURCES:

•Human blastocysts, IVF procedures

•iPSCs

DIAGNOSES:

•Many

•Experimental stage, few clinical trials only

•Geron trial 2009 on spinal chord injury

EX VIVO MANIPULATED IMMUNE CELLS

63

SOURCES:

•BM, PB

ADOPTIVE/ SELECTIVE IMMUNOTHERAPY

•dendritic cells (DCs),

•cytotoxic t lymphocytes (CTLs)

•natural killer cells (NKs)

DIAGNOSES:

• Cancer

• Autoimmune disease

• Several clinical trials going on

IN VITRO CULTURE OF BLOOD CELLS

64

SOURCES:

•BM, PB, UBC

RED CELLS

PLATELETS

IMMUNE CELLS

•dendritic cells (DCs),

•cytotoxic t lymphocytes (CTLs)

•natural killer cells (NKs)

DIAGNOSES:

•

Contents


2

3 SAFETY: NAT, pathogen inactivation

4


6

7

R&D PROJECTS & EXPERIENCES – BTC Ljubljana



67

68

69

Microscope in hood

Introduction to Human Embryonic Stem Cell Culture MethodsCopyright @ 2003 WiCell Research Institute, Inc

A static enclosure, dedicated to removing colonies, is essential. A dissecting scope will fit inside

nicely after the creation of an opening, which theeyepieces can fit through.

70

71

72

73

RESEARCH PROGRAMME 2008-2011

1. TITLE: HUMAN STEM CELLS – ADVANCED

CELL THERAPY

2. LEADER: Ass.Prof.Primož Rožman, MD, PhD

3. PERFORMERS

0311 Blood Transfusion Centre of Slovenia, Ljubljana

0312 University Medical Centre, Ljubljana– Department of Haematology (dr. Samo Zver),

– Department of Gastroenterology (prof. dr. Borut Štabuc),

– Department of Surgical Infections (prof. dr. Dragica Smrke)

– Department of Orthopaedic Surgery (dr.Vane Antolič)

7421 EDUCELL Ltd, Cell Therapy Service, Ljubljana

4. ANNUAL RESEARCH HOURS: 3650

74

ESC

New: Pluripotent SC in adult?

D ESC-A

77

ESC-A IN THE OVARIAN ENDOTHELIUM?

Virant Klun,

Rožman et al.

2008

79

89

Clinical study: The healing of hard-to-heal fractures of long bones by growth factors/autologous SCs from BM. (+ Department of Surgical Infections).

Background: platelet GFs induce in SCs an increase of bone formation.

Design: treatment with GFs/ SCs mixture. Prospective study in 20 patients.

Evaluation: The success rate in two groups of patients.

Expected results: A statistically improved healing of fractures in patients receiving the SC/platelet gel grafts.

3. Differentiation of SCs into the bone

90

Platelets: various growth factors are

contained in the platelet gel

91

Composite graft

92

A) Predoperativno

B) Postoperativno – 12 m.

Bones

93

Clinical study: Modulation of immune response in pts. with tubular adenoCa of pancreas. Coll. with Dept. of Gastroenterology

Background: the Tx of allogeneic MNCs (the GVT effect) which are death-programmed in order to prevent the GVHD effect but retain the GVT effect.

Design: 15 x 2 patients, conditioning with CS, appl. f modified allogeneic MNCs.

Expected results: extended OS and quality of lifeGenomics, proteomics, transcriptomics: explane underlyingmechanisms

4. Cellular immune therapy of solid tumours

The idea: to enhance the normal anti-Tu immunity

Tumour specific

regulatory T-cells

REMOVE

GVHDGVL-GVT

TUMOUR

96

Programme team (consortium):

The team has been working together > 10 years

Novel technologies and more than 10 working places created

>180 patients successfully treated using advanced cellular therapies

Collaboration:

Columbia University (3 researchers currently in New York)

Newcastle University (1 researcher)

Imperial College London (prof. Habib)

King´s college London (prof. S.Minger)

Continuity: ARRS

7 ongoing projects

11 closed projects

99

CONCLUSION: TRANSFUSION MEDICINE -

GLOBAL CHANGE OF PARADIGM

1. Provision of blood products, donor recruitment, blood banking

2. Hemotherapy – clinical TM

3. Transplantation, histocompatibility, immunology

4. Other therapies: biotechnology, advanced therapies

(cellular th., gene th., tissue eng.)5. Laboratory diagnostics related to blood transfusion/transplantation

6. Acompanying activities:National registries of bone marrow donors

Biobanking – Umbilical Cord Blood Bank, Tissue Bank, Organ Bank

Prenatal program for HDN prevention

100

FUTURE OF NIGHT SHIFTS

102

University Clinical

Centre, Ljubljana

Dragica Smrke

Borut Gubina

Irma Virant Klun

BTC of Slovenia

Dragoslav DomanovićMiomir KneževićMatjaž Jeras Tadeja DovčMarko StrbadElvira MaličevPolona KlemencMetka KrašnaAna TomšičMojca JežPrimož PoženelUrška Tajnšek

Documents

Transfusion medicine changes of the paradigm