Transfusion-related acute lung injury (TRALI) in an obstetric patient.

Transfusion-related lung injury (TRALI) is the leading cause of mortality following transfusion of blood products. Despite increasing awareness, the condition often remains unrecognised and therefore underreported. A 28-year-old with moderate preeclampsia had a post-partum haemorrhage following emergency caesarean section. Shortly after receiving three units of packed red cells she went into respiratory failure, which progressed to cardiac arrest. She was successfully resuscitated and made a slow but full recovery. Investigation through the National Blood Service confirmed the diagnosis of TRALI. TRALI is an increasingly common life-threatening complication of blood transfusion and should be included in the differential diagnosis of collapse in an obstetric patient who has recently received a blood product transfusion.

Anti-D immunoglobulin treatment for thrombocytopenia associated with primary antibody deficiency.

AIMS: To review our experience of anti-D immunoglobulin for immune thrombocytopenia (ITP) in patients with primary antibody deficiency. METHODS/PATIENTS: A retrospective case notes review of four Rhesus positive patients with ITP and primary antibody deficiency, treated with anti-D. Patients were refractory to steroids and high dose intravenous immunoglobulin (IVIG). Two patients were previously splenectomised. RESULTS: All patients responded to anti-D immunoglobulin. Improved platelet counts were sustained for at least three months. Side effects included a fall in haemoglobin in all cases; one patient required red blood cell transfusion. Two patients had transient neutropenia (< 1 x 10(9)/litre). CONCLUSION: Anti-D immunoglobulin may be an effective treatment for antibody deficiency associated thrombocytopenia, even after splenectomy. Anti-D immunoglobulin may have considerable clinical advantages in this group of patients, where treatments resulting in further immunosuppression are relatively contraindicated.

Aggressive NK cell lymphoma preceded by a ten year history of neutropenia associated with large granular lymphocyte lymphocytosis.

We report a case of aggressive natural killer (NK) cell lymphoma in an 82 year old man who first presented 10 years earlier with neutropenia in association with a large granular lymphocyte (LGL) lymphocytosis. The diagnosis of NK cell lymphoma was made on the basis of morphological and immunological characteristics (CD3-CD56+) found on skin biopsy of one of multiple skin nodules which subsequently developed in association with splenomegaly, thrombocytopenia and continuing neutropenia. In addition there was BM infiltration and a cytogenetic abnormality [add(6)(p25)] was detected. Combination chemotherapy led to an initial clinical response but a relapse occurred shortly afterwards and the patient died 8 months later from infection whilst neutropenic following re-introduction of chemotherapy. Previously reported cases of aggressive NK cell lymphoma have shown a young male predominance with a rapidly progressive clinical course and without evidence of a preceding chronic phase of LGL lymphocytosis and neutropenia.

Deficiency of glycosylphosphatidyl inositol-anchored proteins in patients with aplastic anaemia does not affect response to immunosuppressive therapy.

Deficient expression of glycosylphosphatidyl inositol (GPI)-anchored proteins in aplastic anaemia (AA) patients has previously been reported to be associated with a poor response to immunosuppressive (IS) therapy. Here we report the response to IS therapy of 111 patients with AA and correlate this with GPI-anchored protein expression on peripheral blood cells by flow cytometry. A GPI-anchored protein deficient population was identified in 15% (17/111) of patients with AA who had a negative Ham's test and no laboratory evidence of haemolysis. Patients were treated with antilymphocyte globulin and/or cyclosporin A, or oxymetholone. Bone marrow transplantation was performed in 12 patients, seven of whom had not responded to IS therapy. In patients tested for CPI-anchored protein expression prior to IS therapy there was no difference in response rate to IS therapy between AA patients with a GPI-anchored protein deficiency and those with normal GPI-anchored protein expression (50% response rate versus 75%, respectively). Survival in these two groups was similar at 90% with follow-up over 140 months from diagnosis. Eight of the 17 AA patients who developed a GPI-anchored protein-deficient population later went on to develop a positive Ham's test. From this study we demonstrate a lower incidence of GPI-anchored protein deficiency in AA patients compared with previous reports. In addition we have shown that the presence of a GPI-anchored protein-deficient cell population in patients with AA who have a negative Ham's test is not a poor prognostic factor in terms of response and survival after IS therapy.

Familial acute myeloid leukaemia: four male members of a single family over three consecutive generations exhibiting anticipation.

A kindred with familial acute myeloid leukaemia (AML) is described displaying the concept of anticipation which refers to increased disease severity and/or earlier age at onset with succeeding generations. The affected individual in the first generation was the father (aged 34 years at diagnosis) of two affected members of a single sibship (aged 24 and 25 years). The fourth patient is the son (aged 4 years) of one of these brothers in the second generation. The AML was of different subtypes and a karyotypic abnormality (del[6q]) was detected in one of three patients. The first-generation patient died during chemotherapy; the remaining three patients are in complete remission after chemotherapy and autologous bone marrow transplantation.

Fatal autoimmune pancytopenia following bone marrow transplantation for aplastic anaemia.

We report a case of autoimmune pancytopenia 10 months after allogeneic bone marrow transplantation (BMT) for severe aplastic anaemia (SAA). The autoimmune haemolytic anaemia (AIHA) and immune thrombocytopenic purpura (ITP) were refractory to conventional immunosuppressive therapy which included steroids, azathioprine, vincristine and intravenous immunoglobulin. Splenectomy led to a recovery of the thrombocytopenia but the haemolysis continued despite further immunosuppressive therapy. Four months after the onset of haemolysis granulocyte-specific antibodies were detected. The patient subsequently received total lymph node irradiation (TLI) with a peripheral blood stem cell transplant (PBSCT) from his original donor, but died 9 days later from cerebral aspergillosis. The severe nature of autoimmune cytopenias and their lack of response to conventional treatment following allogeneic BMT is discussed further.

Interleukin-4 enhances the survival of severe combined immunodeficient mice engrafted with human B-cell precursor leukemia.

Human interleukin-4 (huIL-4) has been shown to inhibit the growth in vitro of cells from patients with acute lymphoblastic leukemia (ALL). With the aim of determining whether this cytokine might be useful in the treatment of patients with ALL, the effects of huIL-4 on human B-cell precursor ALL engrafted in severe combined immunodeficient (SCID) mice were examined. The inhibition of [3H] thymidine uptake of primary ALL cells by huIL-4 was maintained following engraftment and passage of leukemia in SCID mice. Five of seven xenograft leukemias showed significant inhibition in vitro by huIL-4 at concentrations as low as 0.5 ng/mL; furthermore, huIL-4 counteracted the proliferative effects of IL-7. When used to treat two human leukemias engrafted in SCID mice, huIL-4 200 microgram/kg/d, as a continuous 14-day subcutaneous infusion, suppressed the appearance of circulating lymphoblasts and extended survival of mice by 39% and 108%, respectively, the first demonstration of IL-4 activity against human leukemia in vivo. The mean steady-state huIL-4 level in mouse plasma during the infusion was 1.46 ng/mL (SEM +/- 0.14 ng/mL), which was similar to concentrations found to be effective in vitro. ALL cells obtained from mice relapsing after huIL-4 treatment continued to show inhibition by the cytokine in vitro. These data suggest that IL-4 may be useful in the treatment of patients with ALL.

Genes and gene therapy--prospects and problems in the blood.

Advances in recombinant DNA technology and in the understanding of the molecular biology of human disease have made the development of human gene therapy a realistic goal. A conference held at The Royal College of Physicians on 29 September 1994 provided fascinating insights into prospects and problems of gene therapy in haematological practice.

Deletions and rearrangement of CDKN2 in lymphoid malignancy.

Recurrent abnormalities of the short arm of chromosome 9, including translocations and interstitial deletions, have been reported in both leukemia and lymphoma. The pathologic consequences of these abnormalities remain unknown. The cyclin-dependent kinase 4 inhibitor (CDKN2) gene, which maps to 9p21, has been implicated by the finding of a high frequency of biallelic deletions in leukemic cell lines. We have determined the incidence of structural abnormalities affecting CDKN2 by DNA blot in a panel of 231 cases of leukemia and lymphoma and 66 cell lines derived from patients with lymphoid malignancies with defined cytogenetic abnormalities. Structural alterations of CDKN2 were seen in 20 (8.3%) of all fresh cases and 10 (15.1%) of all cell lines. Biallelic CDKN2 deletions were seen in 11 of 53 (21%) cases of B-cell precursor acute lymphoblastic leukemia (BCP-ALL). There was no association with any particular cytogenetic abnormality. Biallelic deletions were also found in high-grade and transformed non-Hodgkin's lymphoma (NHL) of both B- and T-cell lineages. In two cases of transformed NHL, analysis of sequential samples showed loss of CDKN2 with transformation. Neither deletions nor rearrangements of the CDKN2 gene were seen in any of the 119 leukemias of mature B or T cells analyzed. Biallelic deletions of CDKN2 were observed in 6 of 13 NHL cell lines. Three of the 6 cases had undergone transformation from low- to high-grade disease: in 2 of these cases it was possible to show that the CDKN2 deletions were present in fresh material from the patient and were therefore not an artifact of in vitro culture. Rearrangements of CDKN2 were seen in 2 cases (4%) of BCP-ALL, in 1 case of B-NHL, and in 1 Burkitt's lymphoma cell line and suggest the presence of a "hot spot" for recombination in the vicinity of the CDKN2 gene. These data indicate that the loss of CDKN2 expression may be involved in the pathogenesis of a subset of BCP-ALL, some high-grade NHL, and in the transformation of NHL from low- to high-grade disease. CDKN2 deletions and rearrangements occurred in the absence of detectable cytogenetic changes of chromosome 9p in 25 of 30 (83%) cases. Finally, of 10 cases of BCP-ALL that produced overt, transplantable leukemia in mice with severe combined immunodeficiency (SCID), seven showed biallelic CDKN2 deletions. In contrast, none of 11 cases that failed to engraft showed biallelic CDKN2 deletions. BCP-ALL cases that lack CDKN2 expression may have a particular propensity to grow in SCID mice.

Growth of primary human acute lymphoblastic and myeloblastic leukemia in SCID mice.

The in vivo growth of 8 human primary acute lymphoblastic (ALL) and 17 primary acute myeloblastic (AML) cell populations was investigated in severe combined immunodeficient (SCID) mice. Bone marrow (BM) or peripheral blood (PB) samples, either fresh or cryopreserved, were implanted i.v. into irradiated SCID mice. The cells from 5/8 patients with ALL resulted in engraftment with systemic proliferation and dissemination leading to morbidity and mortality of the animals within 12-18 weeks from implantation. In contrast, none of the 17 AML samples resulted in sustained engraftment. Four of the 5 engrafted ALL populations have been successfully passaged into fresh recipients and phenotypic and karyotypic characteristics remain unaltered. The data presented indicate that the SCID mouse may be a useful model for studying the pathogenesis of ALL and may also enable the development and investigation of new therapies for this disease.

Graft failure after bone marrow transplantation from unrelated donors using busulphan and cyclophosphamide for conditioning.

Five patients received bone marrow allografts for leukaemia from serologically fully matched unrelated donors. The conditioning regimen was busulphan 16 mg/kg and cyclophosphamide (CY) 120 mg/kg. Engraftment was achieved in only two patients, with primary graft failure occurring in three patients. Two of the three patients with no evidence of myeloid engraftment on day 28 received GM-CSF without response. Both these patients required rescue with cryopreserved autologous material. Three patients remain alive, one died of relapse after a successful allograft and one died with pancytopenia. Although the number of patients is small, we conclude that the BuCY2 regimen with 120 mg/kg CY, while adequate for sibling allografts, may not be sufficiently immunosuppressive to permit consistent engraftment of bone marrow from unrelated donors.

Human Philadelphia chromosome-positive chronic myeloid leukemia: a potential model for antisense therapy.

We have developed an in vivo model of human chronic myeloid leukemia (CML). A peripheral blood (PB) sample of Philadelphia (Ph) chromosome-positive CML cells in lymphoid blast crisis was transplanted intravenously (IV) into sublethally irradiated severe combined immunodeficient (SCID) mice, and this resulted in engraftment with systemic proliferation. Growth of leukemia was monitored by PB cell morphology and by flow cytometric analysis of murine PB cells labelled with an anti-human leukocyte antigen (HLA) monoclonal antibody. Human cells were first detected in the PB at 4 weeks and comprised a mean of 57% of the total nucleated cells in the PB of these mice by 15 weeks. The Ph chromosome was retained and the population has been successfully passaged. BCR/ABL fusion gene expression was detected in a subsequent passage. Experiments are underway to use this in vivo model to assess the antileukemic activity of BCR/ABL antisense oligonucleotides.

Failure of IMVP-16 as second-line treatment for relapsed or refractory high grade non-Hodgkin's lymphoma.

The British National Lymphoma Investigation (BNLI) has assessed the use of an IMVP-16 regimen (ifosfamide, methotrexate, VP-16) in 46 patients with high grade non-Hodgkin's lymphoma (NHL) who on first-line chemotherapy either failed to attain a complete remission or relapsed. Seventeen patients responded to IMVP-16 but only five (11 per cent) went into a complete remission (CR), 12 (26 per cent) had a partial remission (PR) and 29 (63 per cent) showed no response (NR). CR after IMVP-16 has been maintained in only one case (36 months). The results of this study imply that the use of this IMVP-16 protocol as second-line treatment for patients with recurrent high grade NHL is unsuccessful and alternative salvage regimens should be sought.

Growth of primary human acute leukemia in severe combined immunodeficient mice.

Seven populations of human leukaemic cells were implanted i.v. into sublethally irradiated severe combined immunodeficient (scid) mice. Growth of leukaemia was monitored by labelling murine peripheral blood (PB) cells with an anti-HLA monoclonal antibody and flow cytometric analysis. Two of the populations transplanted were fresh acute lymphoblastic leukaemia (ALL) bone marrow (BM) cells which both caused sustained proliferative growth in scid mice. Human cells accounted for up to a mean of 87% of the total nucleated cells (TNC) in the PB of these mice between weeks 12-15. One of these populations was passaged into fresh mice and frank leukaemia was again established. Three populations of cryopreserved acute myeloblastic leukaemia (AML) cells (2 obtained from PB and 1 from BM) and one population of cryopreserved biphenotypic acute leukaemia BM cells, only grew to a maximum of 4% within the 15 week period of the experiment. A cell population from an AML cell line (HL60), however, did engraft and proliferate resulting in a rapid deterioration of these mice between weeks 3-6 when the proportion of human cells accounted for 9% of the TNC in the PB.