Case Ri'ports

British journal of Haematology, 1992, 82

A 1. I ,O c; EN E I C K O N E $1 A K K 0 \\' T K A N S PLANT AT I 0 N F 0K P K I 34 AK Y M Y E LOF I BKOS I S

The literature contains notably scarce information with regard to the use of allogeneic niarrow transplantation as therapy for myelofibrosis. I\'? report the details of a 49-yearold male with primary myelofibrosis who received marrow ablative chemotherapy followed by allogeneic bone marrow transplantation (BMT) as primary therapy. Post-transplant there was no engraftment and after a few months there were signs of autologous recovery. In September 1988 at the age of 46 years. the man had presented complaining of fatigue and night sweats. There was splenomegaly of 6 cm below the left costal margin and hepatomegaly. Blood counts were: haemoglobin 6.9 mniolil. white blood cells 4 . i x 10'' I. granulocytes 2.1 x 10yjl,and platelets 4 7 x 1Oq,4. The blood smear revealed leuco-erythroblastosis with tear-drop poikylocvtosis and platelet polymorphism. A bone marrow biopsy from the iliac crest showed fibrosis. with a marked increase in reticulin. striking clustering of tnegakaryocytes and virtual absence of normal haemopoiesis. As he became more dependent on erythrocyte transfusions and prone to infections it was decided 3 years after diagnosis to perforni BXlT with his -14-year-old HLA identical, MLC negative brother a s the donor. At the time of transplantation his performance status iras relatively good. with a splenomegaly of 7 cm and no hepatomegaly. He was entirely transfusion dependent. leucoplatelets cytes 2 . 3 x 10'' 'I. granulocytes 0 . h x 1O''!I. 3 0 x 10v. The pretransplant conditioning regimen consisted of etoposide 2 x 2 0 mg/kg. cyclophosphamide 2 x h 0 mglkg and fractionated total body irradiation ( 2 x 6 . 0 Gy. lung dose I x 4 . 2 5 Gy). Partial T-cc4l depletion of the harvested bone marrow was performed by E-rosetting with sheep erythrocytes. The graft contained 1 ' 9 0 x 10- nucleated cells per kg and 49.4 x 1O4 E-rosette-positive T-cells per lig. The number of CFli-GI11 was 8.1 x 10' kg. the number of BFI'-E I i . 6 x 1 0 ' 'kg. N o further graft-versus-host prophylaxis was given. After transplantation there ivas a gradual decrease of the splenomegaly. but there rvere no signs of engraftment. Kepeated bone rtiarrcr\v biopsies revealed fibrosis with coii1plete absence of haemopoiesis. On day 48 we performed a second HMT without additional conditioning with whole marrow from the same donor. The graft was performed following Ficoll-lsopaquc separation and contained 7.1 2 x 10: nucleated cells per kg and 1 . 9 8 x 10; T-cells. 2 1 .6 x l o 4 CFt.-GhI and 18.7 x 10' BFI'-E per kg. Again there were no signs of cngraftnient. It appeared that the spleen increased in size. Treatment with the haemopoietic growth factor G-CSF ( 5 I'glkg once daily, subcutaneously) was initiated to stimulate granulopoiesis. Finally. iifter a period of more than 3 months blood cells commenced to recover white cell iso-enzymes and antigen typing of the red cells indicated that these were autologous cells. Blood counts reached values similar to those seen before transplantation and splenomegaly returned to pretransplant magnitude. At the tirne of writing I 1 0 months after the first BMT) the patient is reasonably well but fully transfusion dependent.

Primary myelofibrosis. also known as agnogenic myeloid tnetaplasia. is a chronic myeloproliferative disorder, which mainly occurs after the age of 50. Survival from diagnosis may vary from less than 1 year to more than 10 years. Conventional treatment is based on relief of symptoms. Cytotoxic drugs o r radiotherapy may decrease or eliminate constitutional symptoms such as night sweats or weight loss, decrease the size of the spleen and liver and sometimes lead to improvement of platelet and leucocyte counts and resolution of inarrow fibrosis (Hasselbach, 1990; Weinstein. 1 9 9 1). Recently. Dokal ~t a/ (1989) and Ifrah et a! (1989) have reported the successful treatment of two patients with myelofibrosis using allogeneic BMT. Both showed rapid engraftment of all cell lines probably enhanced by prior splenectorny . According to a n International Bone Marrow Transplant Registry (IBMTK) analysis, BMT has rarely been applied in patients with primary myelofibrosis. A total of 1 3 cases has been registered (M. Bortin, personal communication). The data presented here were obtained from the Statistical Centre of the International Bone Marrow Transplant Kegistry: the analysis has not been reviewed or approved by the Advisory Committee of the IBMTK).Of these, eight patients survived between 4 and 71 months after allogeneic BMT. Rajantie et a1 (1 986) showed, in a retrospective study. that marrow fibrosis may adversely affect posttransplant haematopoietic recovery. They hypothesized that marrow obliterated by dense fibrosis could be unable to provide an adequate micro-environment for haematopoietic reconstitution. Alternatively. it may be possible that persisting fibrosis is the indication that the malignant process has not been ablated, and hence that the malignancy itself impairs engraftment. Longmore et a1 ( 1 990) reported their results of transplantation in patients with myelodysplasia and secondary acute non-lymphoblastic leukaemia. In a subgroup with marrow fibrosis it was noteworthy that the only patients with marrow failure after allogeneic BMT had T-cell depletion of the graft. In conclusion. bone marrow transplantation is a therapeutic option for younger patients with primary myelofibrosis. However, failure to engraft is a significant risk factor. It may be necessary to avoid T-cell depletion and to perform splenectomy prior to transplantation as efforts to maximally promote repopulation.

REFERENCES Dokal. 1.. Jones. L.. 1)eenmamodr. M., Lewis. S.M. & Goldman. 1.M. [ 1989) Aliogeneic bnnc marrow trarisplantatioii for primary rnvelofibrosis. British joirrrinl o j Wic~rrrutoloqg.71, 1 58-1 59. Hasselbach. H. ( 1 990) Idiopathic myelofibrosis: a review. Eirrope~ti loirrnnl of Hnrwritolo.qy. 45, 65-72,

British Journal of Haematology, 1992, 82

Case Reports

Ifrah. N.. Coudembas-Pain, M., Hunault. M.. Saint-Andrc, J.P., Foussard, C;. & Boassau, M. (1989) Allogeneic bone marrow transplantation for primary myelofibrosis. British Journal ojHrccwiatology. 7 3 , 575-576. Longmore. G.. (;tiinan. E.C.. Weinstein. H.J., Gelba. R.D., Rappeport, 1.M. & Antin. J.H. (1990) Bone marrow transplantation for myelodysplastic and secondary acute non-lymphoblastic leukemia. ]ournrtl o/Clinical Oncology. 8, 1707-1 714.


Rajantie, J.. Sale, G.E.. Deeg. H.J.. Amos. D.. Applebaum. F.. Storb. R.. Clift, R.A. & Buckner. C.D. ( I 986) Adverse effect ofsevere mm-ow fibrosis on hematologic recovery after chernoradiotherapp and allogeneic hone marrow transplantation. Hluod. 67, 169 3-1 697. Weinstein, T.M. ( 1 991 ) Idiopathic myelofibrosis: historical rwiew, diagnosis and management. Blood Rc,vicws. 5. 98- 104.


Polycythaemia vera (PV) shares several features with the other non-leukaemic myeloproliferative disorders (MPD), myelofibrosis and essential thrombocythaemia, both clinically and in vitro (Adams et a/, 1988). These include clonal haemopoiesis and the growth of apparent erythropoietin (Epo)-independent erythroid progenitor cells in clonogenic assay (endogenous erythroid colonies: EEC). A 6 7-year-old man presented with a raised haemoglobin in 199 1. Further clinical and laboratory investigations confirmed a diagnosis of polycythaemia vera. Cytogenetic analysis of peripheral blood showed a normal male karyotype and the presence of two hyperdiploid clones with trisomy 8 in addition to other chromosomal abnormalities: 47, XY, 8, + 9 , -12,t(6: 12)/48,XY. f 8 , f 9 , -12,t(6; 12).del(l2q). Brief details of this patient have been published previously (Price et al, 19921. Perrpheral blood mononuclear cells at presentation were cultured in a standard methylcellulose assay for erythrnid progenitor cells. Some of the cultures were initiated with 5 units/ml of Epo while the rest were grown without Epo. After 1 4 d incubation individual erythroid colonies were removed using a micropipette, suspended in phosphate buffered saline (PBS) and cytocentrifuged onto glass microscope slides. The slides were either fixed in methanol and Giemsa stained to confirm morphological cell type or air dried and stored wrapped in foil at - 20°C for later simultaneous immunophenotypic and genotypic analysis


(Price et al. 1992). These latter slides were fixed in acetone and stained with an anti-glycophorin A antibody (R10: a gift from Dr P. Edwards) using a previously described three-stage alkaline phosphatase anti-alkaline phosphatase method (APAAP)(Cordell et al, 1984), with Fast Red as the substrate. Fluorescence in-situ hybridization (FISH) with a chromosome 8 specific alpha-satellite probe (Oncor Science. Gaithersburg, Md.), was performed on cytocentrifuged cells (some immunophenotyped, see above) as described previously (Price et a/, 1992). Cultures grown in the presence of Epo produced 29 1 f24 erythroid colonies (per 2 x 105 cells plate&SE). while cultures without Epo grew 1 0 4 f 9 colonies. The erythroid nature of these colonies was established by their characteristic morphology and red colour in the culture plates and by Gienisa staining or, in some cases, immunophenotyping. I n our laboratory we have not previously observed EEC generation in peripheral blood MNC cultures from normal subjects using this culture system. 3 5 colonies were picked from the culture plates, cytocentrifuged and analysed by FISH (20/35 were initially irnmunophenotyped). A median of 44 cells/colony (range 7-238) were scored for the number of chromosome 8 signals. 2 0 of these colonies had been grown with Epo and 1 5 without. Of the 2 0 Epo-stimulated colonies, six (30%)had the normal diploid complement of chromosome 8 (two signals) and 14

Fig 1 . Simultaneous immunophenotyping (anti-glycophorin A antibody: red) with in-situ hybridixation (green) for chromosome 8: (A) cell from Epo-stimulated colony showing trisomy 8;(B) cell from Epo-independent colony showing two signals (normal).

Allogeneic bone marrow transplantation for primary myelofibrosis.

772 Case Ri'ports British journal of Haematology, 1992, 82 A 1. I ,O c; EN E I C K O N E $1 A K K 0 \\' T K A N S PLANT AT I 0 N F 0K P K I 34 AK Y...
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