PLATELET REFRACTORINESS

Defined as.
Consistently inadequate response to platelet transfusions Assessment of efficacy of platelet transfusions Corrected Count Increment(CCI) in m2/l=
Post transfusion count-pre transfusion count Number of platelets transfused
RDP=5.51010 SDP=31011

Body surface area m2

1011

 Surface area-dubois formula .007184xht in cm

.725

wt In kg.425

Platelet increments :other formulas Platelet increments :other formulas

MPI =

Platelets Transfused x Sequestration factor

Blood Volumex1000

Adjusts for sequestration & Blood Volume

PPR =

Platelet Increment MPI

100%

CCI= Corrected Count Increment ;; MPI = Maximal Platelet Increment; PPR = Percent Platelet CCI= Corrected Count Increment MPI = Maximal Platelet Increment; PPR = Percent Platelet

Relation between CCI and PPR

PPR=.0026XCCI/F %
CCI 1000 5000 10000 15000 20000 25000 PPRs 3.1 20 39 59 78 101

Minimum requirement for a successful transfusion PPR

> 30% at 1 h > 20% at 2024 h Upto 66% recovery possible

CCI
7500 at 1 h 4500 at 2024 h

Platelet refractoriness
Two consecutive platelet transfusions lead to 1 hr post transfusion CCI<5000 platelets 3 platelet transfusions(not necessarily consecutive)over a 2 week period with inadequate post transfusion counts

WHY.?

Platelet refractoriness-Causes
Non immune causes
Active bleeding (even subclinical) Fever Sepsis Splenomegaly Disseminated intravascular coagulation Veno-occlusive disease Drug-induced thrombocytopenia (ie, amphotericin B, vancomycin) Thrombotic thrombocytopenic purpura Bone marrow and early post-stem cell transplantation Platelet age (>3 d) and poorly stored platelet concentrates

Platelet refractorinessCauses
IMMUNE
Alloantibodies(alloimmunization) Autoantibodies Drug induced antibodies Immune complexes

 Incidence
20-70% of patients who receive multiple transfusions 60-65% non immune causes

Trend in platelet count increment can predict the cause

PLATELET ALLOIMMUNISATION
Formation of antibodies directed to foreign antigens on the surface of platelets May /may not cause platelet refractoriness

Prevalence and Incidence of platelet alloimmunisation

Variable incidence : upto 50%* (multi transfused patients) Kiefel, et al, Transfusion, 2001: 766-70.
252 patients with solid tumor and hematological malignancies, heavily transfused, platelet refractory. 42% anti-HLA antibodies. 8% anti-HPA antibodies.

TRAP trial: 27% developed LCT antibodies.

Risk factors
High number of transfusions? Disease &immunosuppression
Hematological malignancies: Blood, 1987; pp. 1727-9: incidence of LCT in AML 44% vs. ALL 18%. Contribution of steroids? Blood, 1981: pp. 122-8: Heavily transfused patients, examine for LCT. Acute leukemia: 20/65.
Lymphoma: 5/19. Sarcoma: 2/16. Aplastic anemia: 7/8.

Dutcher, et al, Blood, 1981: 1007-11.

Product used

RDP/SDP Leuco reduction

PLATELET ANTIGENS
IX IIIa V IIb Ia IbB Ibalpha

Platelet
ABH IIa

CD109 B2 microglobulin HLA I

Platelet Specific Antigens : HPA (Pl, Bak, Pen etc) HLA Class I - 50- 180.000 molecules ABO Antigens- ABH, Le, Ii , P

A N

A N TI H P

T AN LA IH

Antibodies causing platelet alloimmunisation

Anti HLA antibodies
Most common 18-50% Immunizing Source mainly WBCs not platelets Leuko reduction helps to prevent

Test to detect
LCT test-lymphocytotoxicity assay Panel reactive antibodies >20%
SPRCA

Platelet Surface: Class I HLA Antigens only

WBC Surface: Class II HLA Antigens

Torloni MD - 2002

Antibodies causing platelet alloimmunisation-contd..

Anti platelet antibodies
HPA-1a Rare ,<10% HPA-1b,2a etc

Tests

Phase I,II,III Immunoflourescence/flow cytometry-very sensitive ,not platelet specific MAIPA-Widely used-platelet specific Molecular SSP PCR Typing

ABO system antibodies

ABO system antibodies Major mismatch
Significant in CCI reported,still enough to prevent bleed High expressor donor High titre antibody in recipient

Minor mismatch
Circulating immune complexes

But inventory usually inadequate

PREVENTION
SDP
Decreases the number of donor exposures Process leucoreduction

Leucocyte depletion of platelet productsmost practical

Platelets lack class II HLA Ag WBC responsible for I0 sensitisation

UV irradiation- prevents the interaction of APCs with recipient T lymphocytes

Other effects of platelet alloimmunisation

NAIT PTP-Post transfusion Purpura
PLA-1 mediated usually One week after transfusion Sudden self limiting h/o transfusion/pregnancy Glucocorticoid+plasma exchange/IvIg Future RBC transfusions from HPA-1 a negative donors Washed RBC offer some protection

Diagnosis and management

Non immune
Correct the underlying cause Try dose Correction of anemia

 Fresh platelets
Less than 48 hrs Storage decreases function May be useful in some non immune causessepsis,GVHD,amphotericin B

ABO MATCHED PLATELETS

Easier to obtain comparatively Trial is worth before switching to special products

 Lymphocytotoxicity Test Panel Reactive Antibody level-% of panel cells to which recipient has formed antibodies <20% non immune? Anti HPA antibodies?
Review clinically

platelet antibody screening

>20%

 Alloimmunized patient:
HLA-matched platelets Mismatched platelets (CREGS) HLA-Antigen-negative platelets HPA-negative platelets Platelet crossmatch IVIG&others

 HLA Matched platelet

90% refractory cases benefit BJH, 1987; 115-21 Main problem Donor pool 3000-10000

CREGs
American Journal of Hematology, 1977: pp. 219-26. Cross-reactive groups (CREGS):

Platelets mismatched at 1 or 2 cross-reactive HLA antigens produced increments similar to perfect matches. Provides 10 times as many prospective donors.

PRA<60%,One antigen mismatch

Fortunately ,most antibodies spontanously disappear

 HLA A&HLA B match important Other absent/weak Closest match within time and donor availability Should be irradiated HLA typing of recipient better done in advance before leukopenic phase

ASP method Identify the specificity of the antibody, Select compatible donors Allows selection of many more donors Difficulty
broad specificities of antibodies

Not responding still?

Platelet cross match
Radio/flourescent/enzyme linked antiglobulin test Solid Phase Red Cell Adhesion Assay Flow cytometry

Disadvantage
Heavily alloimmunised(PRA>80%)difficult to find compatible

 PIFT Pretreat with paraformaldehyde

Diminished uptake of aggregated IgG Swelling and expulsion of platelet Ig

Incubate with serum Wash Incubate with antiglobulin labelled with fluorescein isothiocyanate Wash and view under fluorescent microscope Non speific reaction less Can use polyspecific Relatively less sensitive FCIT

 ELISA peroxidase Solid phase RED CELL ADHERENCE ASSAY

Platelets fixed to well of microplate Incubated with serum ,wash Red cells coated with anti IgG added Pellet if negative,coat if positive

MAIPA

Diagnosis and Management

IVIG:
Blood, 1990; 313-16: 400 mg/kg for 5 days. Incompatible platelet donor before and after treatment. Improved 1-6 hour, not 24 hour. American Journal of Hematology, 1991; 15-23: Post transfusion increment increased. Best in patients with PRA<85%. Not as effective as HLA matched Very high dose of IvIg-6g/kg
Seigler et al AmJH1991

Frozen preservation Autologous Much useful in oncology Only effective and reliable therapy to those refractory even to HLA matched DMSO better than glycerol in preserving the function

Protein A column Immune absorption Plasma exchange Cyclosporin A ATG Vinblastin loaded platelet transfusions EACA

 Non responsive to special products??? 3 unsuccessful trials of each special product CPI Continuous platelet infusion 3 RDP/1 SPLIT SDP Q4H Only as therapeutic

Hopefully..
Lyophilised platelets Infusible platelet membranes Thromboerythrocytes Thrombospheres

Refractory to pooled RDP

ABO compatible platelets

Fresh platelets

Platelet antibody tests

-VE

Review clinical factors/dru gs

SELECT COMPATIBLE DONORS BY HLA matching HLA corresponding Ag negative Platelet cross matching

Thank you