Removal of persisting isohaemagglutinins with Ig-Therasorb® immunoadsorption after major ABO-incompatible non-myeloablative allogeneic haematopoietic stem cell transplantation

Werner Rabitsch1, Paul Knöbl2, Hildegard Greinix1, Erika Prinz1, Peter Kalhs1, Walter H. Hörl3 and Kurt Derfler3

1Department of Medicine I, Bone Marrow Transplantation Unit, 2Department of Haematology and Haemostaseology and 3Department of Medicine III, Division of Nephrology and Dialysis, University of Vienna, Vienna, Austria

Correspondence and offprint requests to: Dr Werner Rabitsch, Department of Medicine 1, Bone Marrow Transplantation Unit, Währinger Gürtel 18-20, A-1090 Vienna, Austria. Email: werner.rabitsch{at}akh-wien.ac.at



   Abstract
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Background. Major ABO-incompatibility can be associated with haemolysis, prolonged red cell aplasia (PRCA) and higher peri-transplant mortality resulting from organ toxicity after conventional and non-myeloablative allogeneic haematopoietic stem cell transplantation. Different therapeutic strategies have been developed to allow erythroid reconstitution in these patients.

Methods. We present three patients, who developed PRCA after non-myeloablative allogeneic haematological stem cell transplantation for haematological malignancies. The patients were treated with Ig-Therasorb® immunoadsorption (five treatments per week) to remove persisting incompatible isohaemagglutinins.

Results. Two patients became transfusion independent after 12 and 14 treatments. In one patient, however, no reduction of the isohaemagglutinin titres could be observed after 25 treatments, probably due to persistence of his underlying disease.

Conclusions. Although Ig-Therasorb® immunoadsorption was effective in only two patients, it seems to be a promising therapeutic option for patients with PRCA after allogeneic non-myeloablative haematological stem cell transplantation.

Keywords: blood component removal; blood group incompatibility; haematopoetic stem cell transplantation; red cell aplasia



   Introduction
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Non-myeloablative allogeneic haematopoietic stem cell transplantation (HSCT) seems to be a curative therapeutic option in various haematological diseases [1]. ABO blood group incompatibility between donor and recipient occurs in 30–40% of patients undergoing HSCT [2], as ABO blood groups are inherited independently from human leucocyte antigens [3]. Major ABO-incompatible allogeneic HSCT recipients may experience delayed red cell recovery because of persistent host isohaemagglutinins that suppress donor’s red cell haematopoiesis. Different methods have been used for treatment of this complication after conventional allogeneic HSCT: transfusion of donor-type red cells [4]; removal of ABO antibodies by large-volume plasmapheresis prior or after transplantation [5]; removal of anti-A and anti-B antibodies by immunoadsorption on immobilized blood group antigens [6], erythropoietin [7,8], treatment with rituximab [9] and immunosuppression with glucocorticoids or anti-thymocyte globulin [10].

Persisting incompatible isohaemagglutinins may also occur after non-myeloablative HSCT. Because non-myeloablative HSCT is a new therapy option for various haematological malignancies, no data exists about the treatment of persisting isohaemagglutinins in these patients.

Ig-Therasorb® immunoadsorption is an established method for the treatment of various haematologic and immunologic diseases [11]. We report our experience with Ig-Therasorb® immunoadsorption for the treatment of three patients with prolonged red cell aplasia (PRCA) due to persisting isohaemagglutinins after non-myeloablative HSCT.



   Subjects and methods
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Patients
Three male patients, median age 46 years (range 28–53 years), with PRCA after major ABO-incompatible non-myeloablative HSCT were studied. One patient had received transplantation for multiple myeloma, one for myelodysplastic syndrome and one for non-Hodgkin’s lymphoma. Stem cell donors were HLA-identical siblings in all patients. Two patients received a combined protocol consisting of autologous peripheral blood stem cell transplantation for disease debulking followed by non-myeloablative HSCT. The patient with myelodysplastic syndrome received non-myeloablative HSCT alone. Immunosuppressive conditioning regimen consisted of fludarabine and a single dose of total body irradiation (2 Gy). For graft-versus-host disease prophylaxis cyclosporine and mycophenolate mofetyl were given. In all recipients blood group was O, donor blood group was A in two and B in one case. Blood transfusions were performed when necessary (at haemoglobin values <8.0 g/dl) with packed red blood cells of blood group O.

Blood group, Rhesus typing and haemagglutinin titres
Rhesus typing and identification of complete and incomplete red cell alloantibodies and direct antiglobuline testing were performed by standard methods [11,12]. Commercial reagents (DiaMed-ID system, Cressiesur-Morat, Switzerland) were used according to the manufacturer’s instructions. Titres of anti-A and anti-B haemagglutinins were determined before the conditioning regimen, then weekly until discharge, and thereafter biweekly until stable red cell engraftment had occurred. Anti-A and anti-B haemagglutinins were determined by titrating the patient’s sera against suitable RBC’s (in saline, LISS, and in an indirect antiglobulin test) [12]. To document any treatment benefit of immunoadsorption, a relative haemagglutinine score was evaluated by using the DiaMed-ID microtyping system (results are given as strength of agglutination, rated 0–4) [13].

PRCA
PRCA was defined as reticuolocytopenia <30 g/l that lasted for >60 days after non-myeloablative HSCT, absence of RBC precursors in the bone marrow, and dependence on RBC transfusions after exclusion of RBC alloantibodies, haemolysis, viral or bacterial infection or relapse in patients with myeloid and megakaryocytic regeneration after non-myeloablative HSCT.

Immunoadsorption
Ig-Therasorb® immunoadsorption was performed on Sepharose-coupled polyclonal sheep antibodies against human immunoglobulins as described previously [14], in processing 7000 ml of plasma per treatment [15]. Five treatments per week were performed during the first 2 weeks, then treatment frequency was reduced to three or four times a week until incompatible isohaemagglutinins were no longer detectable and a significant rise of haemoglobin and reticulocyte counts was observed. No immunoglobuline concentrates were substituted during treatment.



   Results
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
All patients studied had sufficient engraftment of neutrophil cells and platelets after non-myeloablative HSCT and a mixed chimerism could be demonstrated on days 28 and 60 by fluorescence in situ hybridization. Red cell reconstitution did not occur until day 60. Bone marrow aspiration showed normal myeloid and megakaryocytic lineage with absent erythroid cells. All patients fulfilled the criteria of PRCA at day 60 after non-myeloablative HSCT. Erythropoietin was given as initial treatment at a dose of 10 000 IU subcutaneously three times a week without beneficial effect. Erythropoietin treatment was continued during immunoadsorption and was terminated when a significant rise in reticulocytes was observed. Two patients were primarily treated with Ig-Therasorb® immunoadsorption, one patient received pre-treatment with five cycles of standard plasmapheresis. Ig-Therasorb® was started on days 59, 79 and 88 after non-myeloablative HSCT. IgG levels were significantly reduced by immunoadsorption in each patient (Patient 1: 635 to <195 mg/dl; Patient 2: 1270 to <195 mg/dl; Patient 3: 1290–276 mg/dl), as well as the IgM levels (Patient 1: 169–92 mg/dl; Patient 2: 212 to <30 mg/dl; Patient 3: 222 to <30 mg/dl), by a single apheresis treatment. In parallel, a decline in the isoagglutinine titres was observed. In two patients the persisting isohaemagglutinins could be completely removed after 12 and 14 treatments (Figure 1, Patients 1 and 2). Haemoglobin levels and reticulocyte counts increased into the normal range without the need for further blood transfusions. During immunoadsorption patients were in need of packed red blood cells transfusion (Patient 1: four packed red blood cells; Patient 2: six packed red blood cells; Patient 3: six packed red blood cells). In the one patient with multiple myeloma no reduction of the isohaemagglutinin titres could be observed after five cycles of standard plasmapheresis and 25 treatments of immunoadsorption (Figure 1, Patient 3). Further evaluation demonstrated that the IgG-paraprotein of the patient was still demonstrable and the myeloma was not eradicated. Ig-Therasorb® treatment was stopped, and immunosuppression was tapered to induce a graft-versus-tumor effect. The patient developed extensive chronic graft-versus-host disease of skin, liver and gut, but the isohaemagglutinins were cleared and erythropoiesis restored. Figure 1 shows the time courses of all patients. Ig-Therasorb® immunoadsorption treatment was well tolerated by all patients, and no side effects occurred.



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Fig. 1. Time courses of three patients with PRCA after ABO incompatible non-myeloablative stem cell transplantation for haematological neoplasia. Left y-axis: relative haemagglutinine score; right y-axis: reticulocyte count (g/l); x-axis: months after stem cell transplantation; open arrows: plasmapheresis; solid arrows: Ig-Therasorb® immunoadsorption; solid lines: incompatible isohaemagglutinine titres; dotted lines: reticulocyte counts.

 


   Discussion
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Non-myeloablative allogeneic HSCT can induce durable complete chimerism and sustained remission in patients with haematological diseases [1]. However, immunologic problems, such as persisting isohaemagglutinins after major ABO-incompatibility, may be similar to that of conventional myeloablative HSCT. Attempts to remove high titres of persisting isohaemagglutinins prior or immediately after HSCT can cause several problems due to technical, logistic or infection reasons, or may be badly tolerated: infusion of donor-type red blood cells despite still measurable isohaemagglutinins can lead to haemolysis with considerable side effects; adsorption of whole blood or plasma on immobilized blood group antigens is an experimental procedure available only in single institutions. As the incidence of severe PRCA is relatively low and the majority of patients are able to clear the isohaemagglutinins spontaneously [3], a prophylactic treatment of all incompatible HSCT recipients is not justified. Our patients did not respond to erythopoietin and still had high titres of incompatible isohaemagglutinins. In contrast to standard plasmapheresis immunoadsorption allows nearly complete clearance of circulating immunoglobulins of all types and subtypes (Ig1–Ig4), without the concomitant substitution of fresh frozen plasma or albumin solutions. These facts have been recently demonstrated [15]. Therefore, we attempted to remove the isohaemagglutinins with a device that we have already used in patients after allogeneic HSCT and which was known to be free of severe side effects. Ig-Therasorb® immunoadsorption was able to remove the antibodies and to restore erythopoiesis in two of the three patients, and to get them transfusion independent. The method failed in the patient with multiple myeloma. We can only speculate about possible reasons. It is not clear whether the persisting paraprotein may have influenced immunogobuline binding on the adsorption columns, whether the persisting myeloma cells have suppressed erythroid engraftment, or whether the intensity of therapies was too low to remove all antibodies. However, the response on graft-versus-host disease after tapering the immunosuppression suggests that some kind of immunologic phenomenon may have influenced erythopoiesis in this patient.

In our opinion immunoadsorption alone is only a symptomatic treatment, which cannot cause a complete sustaining elimination of the native ABO antibodies. We think that the obtained slight reduction of the antibody titres enables the recovery of the donor’s haematopoiesis and a complete elimination of the isohaemagglutinins occurs by the excess of antigens due to the produced donor-type red blood cells. This hypothesis explains the differences in the course of anti-A and anti-B titres in Patient 1. The complete elimination of anti-A is mainly caused by the regenerating donor-type A cells.

Because of the limited number of patients no data on comparison of standard plasmapheresis and immunoadsorption for the treatment of patients with persisting isohaemagglutinins due to major ABO-incompatibility are available. However, it seems to be of interest that in Patient 3 both treatment modalities failed to induce donor-haematopoiesis.

In conclusion, Ig-Therasorb® immunoadsorption seems to be a promising method for a rapid, efficient and safe elimination for persisting isohaemagglutinins in selected cases after non-myeloablative haematological stem cell transplantation. As the number of patients treated with this device is small, no evidence-based recommendations can be given at the moment. Because of the high technical expense and the necessity for highly qualified staff, this therapy should be performed in specialized centres only.



   Acknowledgments
 
We gratefully acknowledge the excellent assistance of the staff and the nurses of the apheresis unit, the blood bank and the bone marrow transplantation unit.

Conflict of interest statement. None declared.



   References
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 

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Received for publication: 17. 2.03
Accepted in revised form: 16. 5.03





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