Diagnosis and treatment of patients refractory to platelet transfusions

Blood Reviews (1998) 12,215-221 0 1998 Harcourt Brace & Co. Ltd

Consensus conference on platelet transfusion

Diagnosis and treatment of patients refractory to platelet transfusions

M. Contreras

platelet doses on average (David Linch, personal communication). Most patients will fare well with random donor platelet concentrates (RDPC) obtained from routine donations of whole blood and containing not less than 5.5~10’” platelets in 50 ml of plasma. Better quality platelet concentrates with less leucocyte contamination can be obtained from routine donations of whole blood that are processed into buffy coats by ‘top and bottom’ systems which are pooled in a closed system using a sterile connecting device (Table 1). The pools are then processed into buffy coat residues and high quality platelet concentrates. Such pools, from four donations, provide an adequate adult dose of 250-300~10~ platelets. Platelet concentrates of an even higher quality can be obtained using third generation cell separators, allowing the collection of one or two doses from a single donor. In general, T-cell depletion, graft-versus-host disease (GVHD), alloimmunization, sepsis, fever, disseminated intravascular coagulation (d.i.c.) and ABO incompatibility of platelet concentrates considerably increase the need for platelet support. Platelets should have as low a content of white cells as possible; febrile reactions, HLA alloimmunization and immunological refractoriness to the transfusion of RDPCs are correlated with the degree of white cell contamination in transfused components. There is a leucocyte threshold for immune-mediated adverse effects which varies depending on the effect and on the individual patient; for example, the level of white cell contamination that leads to HLA immunization is significantly lower than the level that induces febrile transfusion reactions.

INTRODUCTION

There are two main indications for platelet transfusions: (i) therapeutic, for actual bleeding, and (ii) prophylactic, for patients with bone marrow suppression or bone marrow failure, when the platelet count falls below a pre-agreed trigger, which used to be 20-30x109/L, but which is definitely decreasing in recent years. In large cities with large numbers of hospitals and advanced medicine, most of the platelets produced are used prophylactically on a temporary basis. The provision of platelet supportive therapy is essential for the successful outcome of bone marrow transplantation (BMT), peripheral blood stem cell transplantation (PBSCT) and cord blood transplantation. The majority of recipients of BMT need platelets for a minimum period of 2-3 weeks, but in a large proportion of patients this period is prolonged to 5 or more weeks. On the other hand, recipients of PBSCT seem to require platelet support in smaller quantity and for a shorter time, often l-2 weeks. For example, at University College Hospital in London, UK, recipients of autologous BMT require, on average, 20-30 adult platelet doses (300~10~ platelets/ dose) for support during the pancytopenic period whereas recipients of PBSCT require only four adult

Professor Marcela Co&eras MD FRCPath FRCPEdin, Professor of Transfusion Medicine, Royal Free and UCL Medical School, Executive Director. London and South East Zone of the National Blood Service, Colindale Ave, London NW9 SBG; UK

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Table 1 Platelet concentrates prepared at the North London Blood Transfusion Centre Type of PC

PRP*-RDPC Optipool (top and bottom)? Apheresis: PCS+ (Haemonetics) Apheresis: COBE§

Mean plt. contentx109/unit

Mean wbc contentx109/unit

Number tested

76 300 252

0.04 co.05 0.08

446 867 1413

300-700

<0.003

>200

*PRP = platelet concentrate prepared from platelet rich plasma (PRP) obtained from one random blood donation (RDPC). This product is not produced any longer, though it is produced in many countries worldwide. tOptipoo1 = pooled platelet concentrate prepared by the ‘top and bottom’ system from pooled buffy coats obtained from four random blood donations. §COBE = third generation cell separator. Often a ‘double dose’ of platelets can be collected, which can be split into two packs. (data from J.S. Seghatchian.)

DEFINITION Refractoriness to the transfusion of platelets is the failure to achieve the expected platelet increment after two consecutive platelet transfusion episodes. Several formulas have been developed to monitor the effectiveness of platelet transfusions. The platelet count in peripheral blood immediately before and 15, 60 min and/or 24 h after transfusion must be measured. In addition, the body surface area or blood volume of the recipient must be known. One of the accepted standard approaches to estimating post-transfusion platelet increment is to calculate the corrected count increment (CCI) with knowledge of the patient’s body surface area (BSA), as follows: CC1 = platelet increment (109/L) x BSA (m’) 10” platelets transfused The response to platelet transfusions can also be measured as the % recovery of platelets transfused at a defined time post-transfusion: % recovery =

platelet count increment/p1 x blood volume No. platelets transfused

The platelet transfusion outcome is considered a failure if the CC1 at 1 h is ~7.5 or the % recovery at 1 h is ~15-20, or if the CC1 at 20-24 h is ~4.5. Pregnancy and especially transfusion before BMT are major causes of primary HLA alloimmunization and the consequent immunological refractoriness to further platelet transfusions in a proportion of alloimmunized patients; in patients with aplastic anaemia who are transplanted soon after diagnosis, the incidence of refractoriness (and graft rejection) is lower than in those who receive multiple transfusions prior to transplantation. The main factors affecting development of refractoriness due to alloimmunization are: l

The time required for antibody formation: in a previonsly unexposed recipient it is unlikely that

l

l

immunological refractoriness will develop within the first 2 weeks of starting platelet transfusions. The number of transfusions and the white cell load in blood components: as stated above, there appears to be a leucocyte threshold for HLA alloimmunization. It is accepted that primary HLA alloimmunization can be prevented if the white cell load per transfusion episode is ~5 x 106. Immune status of the recipient: immunosuppressed patients on chemotherapy have a significantly lower rate of alloimmunization than patients with aplastic anaemia who are not immunosuppressed. In addition, if patients have been immunized before by pregnancy or transfusion, secondary alloimmunization ensues more rapidly and more easily, requiring lower white cell contamination in blood components.

DIAGNOSIS

AND AETIOLOGY

The diagnosis of immunological refractoriness to platelet transfusions is established after excluding non-alloimmune causes of refractoriness such as: Transfusion of poor quality, non-viable platelets (difficult to prove prospectively). Splenomegaly and hypersplenism Disseminated intravascular coagulation caused by infection, septicaemia or malignancy Infection and fever, in particular CMV infection Drug-related thrombocytopenia Poor platelet transfusion responses in patients treated with Amphotericin B Bone marrow transplantation in itself can lead to increased platelet consumption due to GVHD, CMV infection, formation of autoantibodies, drugs and microangiopathic associated syndromes Platelet autoantibodies

Patients

0 Circulating immune complexes which bind to platelet Fe receptors leading to clearance by the monomuclear phagocytic system. The most common cause of immunological refractoriness to platelet transfusions is alloimmunization to HLA class I antigens. Immune refractoriness can also be due to ABO incompatibility (both major, i.e. group A platelets transfused to 0 recipients and minor: 0 platelets to A recipients), and to plateletspecific antibodies (Table 2). The diagnosis is made using both laboratory and clinical criteria:

Table 2

Platelet

specific

Original 1

name(s)

zw, PlA

HPA-2

Ko, Sib

HPA-3

Bak,

HPA-4

Pen, Yuk

HPA-5

Br, Hc, Zav

HPA-6w

Lek

MANAGEMENT The vast majority of BMT patients supported with platelets from our Service, do not become immunologically refractory to the transfusion of RDPCs. In addition, not all patients with HLA antibodies become

Antigens

Phenotype frequency Caucasians(%)

HPA-la HPA-lb HPA-2a HPA-2b HPA-3a HPA-3b HPA-4a HPA-4b HPA-5a HPA-5b

97.9 26.8 99.3 14.9 87.7 64.1 299.9 co.2 99.2 20.6

HPA-6bw HPA-7a HPA-7b

99.9 0.01

HPA-8bw


HPA-9bw HPA- 1Obw

0.6 -

Tu, Ca

HPA-7w

MO

HPA-SW

Sr

HPA-9w

Max”

HPA-low

La”

Other

antigens

without

svstem

Adapted

in GP* Illa lbcl Ilb Illa la llla Illa Illa

Ilb IIIa

status PLT Nak” Gov” Va= lY” Gro’ Oe” Pe” Vis

*GP: glycoprotein. permission.

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transfusions

antigens

System HPA-

to platelet

specific antibodies (usually by ELISA or flow cytometry). Some centres such as Leiden, in The Netherlands, screen for HLA antibodies and for platelet-specific antibodies as soon as a diagnosis of immunological refractoriness is suspected. IgM platelet autoantibodies should also be investigated since they interfere with the screening tests but do not seem to play a role in the outcome of platelet transfusions. Clinical criteria: if a patient has a poor response to transfused allogeneic platelets, this is demonstrated either by a reduced or absent posttransfusion platelet increment or recovery on at least two occasions, or a shortened posttransfusion platelet survival, or both.

l

o Laboratory: as HLA antibodies are the predominant cause of immunological refractoriness to platelet transfusions, HLA antibody screening by lymphocytotoxicity is used as an indirect method for detecting platelet alloimmunization. If complement-fixing HLA antibodies are not found in a refractory patient, a search should be undertaken for HLA antibodies reacting by the indirect antiglobulin technique (usually by flow cytometry) and for platelet-

refractory

from

Willem

Ouwehand

>98.1 96.0 98.0 Cl.0 Cl.0 dO.01 -

and Paul Metcalfe.

Available

on the website

www.nibsc.ac.uk

V IV CDwl09 Ilb/IlIa W Illa Illa XV lbol with

the authors

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refractory to the transfusion of RDPCs. In fact, reported figures in the literature for HLA immunization in platelet-transfusion dependent patients range from 30 to 70% whilst the incidence of immunological refractoriness ranges from 8 to 40%. Hence, although some centres advocate routine, periodic HLA antibody screening in platelet transfusion-dependent patients, this seems to be an unwarranted, expensive policy. Most patients who are immunologically refractory have HLA antibodies reacting with >60% of the lymphocyte screening panel. Hence, it is clear that HLA alloimmunization is not synonymous with immunological refractoriness. Sometimes, the HLA antibodies are of restricted specificity and then most RDPCs are compatible, and, on other occasions, the HLA antibodies are directed against class I antigens expressed weakly on platelets. Nevertheless, for those patients who become immunologically refractory, two options are available for the provision of platelet concentrates with a satisfactory outcome: (a) HLAmatched or, (b) crossmatch-compatible platelets. (a) In the presence of HLA antibodies, the complexity of the HLA system makes it very difficult to provide multiple HLA-matched donors required to meet the transfusion needs of chronically thrombocytopenic patients. Such platelets can only be obtained from a large (2000-20000) panel of HLA-typed donors and/or patients’ relatives willing to donate single donor platelets (SDPs) by apheresis. These procedures are expensive, time consuming and demanding on donor and operator. SDPs should have no less than 25x10” platelets (Table 1). At our centre in Colindale, not more than 8% of the total number of platelet concentrates requested are for immunologically refractory patients. In our panel of over 2500 HLA-typed donors, those homozygous at the HLA-A and -B loci (e.g. Al-B8/Al-B8) are particularly useful. Some centres select HLA-typed platelet concentrates based on the Leiden system which considers split HLA-A and -B antigens: A match (all HLA split antigens identical for donor and recipient); B 1U (one blank or homozygous antigen in the donor and the other three antigens identical); B2U (two blank or homozygous antigens in the donor, the other two antigens identical for donor and recipient); BlX and B2X (one or two crossreactive antigens within the broad groups and the rest identical between donor and recipient). If no HLA-matched donors are available and the specificity of the HLA antibodies is known, donors compatible with the antibodies are selected, if available. In other centres, acceptable

HLA mismatches are considered in the selection procedure. From the above, and regardless of the method used to provide platelets for immunologically refractory patients, it is obvious that it would be helpful if patients who are likely to become refractory to the transfusion of platelet concentrates were typed for HLA A and B at the time of diagnosis. In some of the centres that provide HLA-matched platelets as first line therapy, a panel of HPA-typed donors is available for the provision for platelet concentrates for that small number of patients who develop HPA antibodies with or without HLA antibodies. (b) In the absence of an HLA-typed panel of donors or of compatible relatives, or if patients become refractory to HLA-matched platelets, plateletspecific antibodies should be investigated again and the ‘platelet crossmatch’ using patient’s serum against donor platelets by ELISA, solid-phase or by flow cytometry will be helpful for the provision of platelet supportive therapy. The occurrence of immunization to platelet specific antigens is unusual in the absence of HLA alloimmunization, despite the fact that platelet-specific antigens have shown occasionally to be immunogenic in leucodepleted blood components. Several centres provide crossmatch-compatible platelets as a first line approach to dealing with all types of immunological refractoriness, (i.e. mostly due to HLA antibodies) thus avoiding the need for a panel of HLA-typed apheresis donors. With the above two platelet transfusion approaches (a and b), successful outcomes in immunological refractoriness range from 50 to 90% of platelet transfusions. It remains to be decided which of the two approaches is more successful at achieving satisfactory platelet increments. When patients become refractory to all available platelets, massive ABO identical platelet transfusions, intravenous IgG (IVIG), plasma exchange, acidtreated platelets to strip HLA antigens, or EACA (epsilon amino caproic acid) can be tried. Of all these alternatives, massive platelet transfusion seems to be the most successful. However, a number of workers have shown that despite the lack of satisfactory posttransfusion platelet increments in immunologically refractory patients, it is possible to activate haemostatic mechanisms after the transfusion of HLAincompatible platelets. PATIENT FOLLOW-UP

Immunological refractoriness to platelet transfusions can disappear in a matter of weeks and patients

Patients

become responsive again to the transfusion of RDPCs. For this reason, it is important to monitor the presence, specificity and potency of HLA antibodies at least monthly throughout the period of platelet transfusion support.

PREVENTION OF HLA ALLOIMMUNIZATION AND IMMUNOLOGICAL REFRACTORINESS Leucodepletion White cell contamination should be reduced to a minimum in cellular blood components transfused to platelet transfusion-dependent patients, since it is alloimmunization to leucocytes that is the major cause of febrile transfusion reactions and of immunological refractoriness to platelet transfusions. It is possible, with the top and bottom systems, to produce platelets with white cell counts significantly lower than those found in platelet concentrates prepared by the ‘PRP’ (platelet rich plasma) method, as can be seen in Table 1. Although it is impossible to prevent primary HLA alloimmunization totally, in view of previous allogeneic exposure by pregnancy or transfusion in many patients, some transplant centres aim at preventing immunological refractoriness altogether by depleting platelet concentrates of white cells either by filtration or by collecting single donor platelet concentrates with third generation cell separators such as the Spectra-COBE with the leucoreduction system (LRS). It has been well documented that alloantigen recognition requires the expression of both class I and class II HLA antigens on the surface of the transfused cells; hence, it appears that dendritic cells, as antigen presenting cells, are particularly important in this respect. As platelets, in contrast to white cells, express only class I but not class II HLA antigens, leucocyte-poor cellular components will prevent immunization to HLA antigens and, consequently, avoid the vast majority of cases of immunological refractoriness. Leucopoor blood components, prepared by centrifugation methods (e.g. Optipress, Compomat) will decrease but not prevent HLA alloimmunization. On the other hand, leucodepleted components, prepared either by filtration or third generation cell separators will prevent HLA alloimmunization. It has been proposed that the threshold for HLA alloimmunization is 5~10~ leucocytes per transfusion episode. UV irradiation of platelets As stated above, a major route of alloimmunization is by means of antigen-presenting cells (APCs). Donor APCs interact with T-helper cells of the transfused recipient to induce the formation of alloantibodies in

refractory

to platelet

transfusions

219

the recipient. UV-irradiation has been documented to interfere with the function of APCs by one of several possible mechanisms: By preventing the APCs from releasing immunoregulatory substances such as interleukins By interfering with a receptor or decreasing the expression of a membrane antigen on the surface of the APCs resulting in an inability of the cell to participate in the immune recognition process By inducing the formation of suppressor cells in the transfused recipient, hence preventing antibody formation. The future The prophylactic platelet transfusion trigger of 20-30x109/L has been lowered by many clinicians in recent years to 10 and even 5x109/L following clinical audit of platelet usage. In addition, the advent of growth factors and stem cell therapy is significantly reducing the period of platelet transfusion dependency in patients requiring marrow ablation therapy. These developments are reducing the clinical problem of immunological refractoriness to platelet transfusions. Finally, if thrombopoietin, and perhaps other growth factors acting earlier in thrombopoiesis, fulfil their promise of decreasing platelet transfusion requirements, the incidence of immunological refractoriness to platelet transfusions should be reduced even further. FURTHER

READING

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Slichter S J, Alloimmune refractoriness to transfused platelets. In Garratty G. Immunobioloa of Transfusion Medicine 1994; pp597-627 M. Dekker. Sugawara S, Abo T, Kumagal K. A simple method to eliminate the antigenicitv of surface class I MHC molecules from the membrane-of viable cells by acid treatment at pH 3: J Immunol Methods 1987; 100: 83-90 van Marwijk Kooy, M. van Prooijen, H.C. Moes, M. et al. Use of leukocyte-depleted platelet concentrates for the prevention of refractoriness and primary HLA alloimmunization: A prospective, randomized trial. Blood 1991; 77: 201 Waters AH, Minchinton RM, Bell R et al. A cross-matching procedure for the selection of donors for alloimmunized patients: Br J Haematol 1981; 48: 59-68 Wu KK. Hoak JC. Koeoke JA. Thoson JS. Selection of compatible platelet donors: a prospective evaluation of three cross-matching techniques: Transfusion 1977; 17: 638-643 Yankee RA, Graff KS: Dowling R, Henderson ES. Selection of unrelated compatible platelet donors by lymphocyte HLAmatching: New Engl J Med 1973; 760-764 ,

1

/