Attempts to improve treatment outcomes in acute myeloid leukemia (AML) in older patients: the results of the United Kingdom Medical Research Council AML11 trial.

In an attempt to improve induction chemotherapy for older patients with acute myeloid leukemia (AML),1314 patients were randomized to 1 of 3 induction treatments for 2 courses of DAT (daunorubicin, cytarabine, and thioguanine) 3 + 10, ADE (daunorubicin, cytarabine, and etoposide) 10 + 3 + 5, or MAC (mitoxantrone-cytarabine). The remission rate in the DAT arm was significantly better than ADE (62% vs 50%; P =.002) or MAC (62% vs 55%; P =.04). This benefit was seen in patients younger and older than 70 years. There were no differences between the induction schedules with respect to overall survival at 5 years (12% vs 8% vs 10%). A total of 226 patients were randomized to receive granulocyte colony-stimulating factor (G-CSF) or placebo as supportive care from day 8 after the end of treatment course 1. The remission rate or survival were not improved by G-CSF, although the median number of days to recover neutrophils to 1.0 x 10(9)/L was reduced by 5 days. Patients who entered remission (n = 371) were randomized to stop after a third course (DAT 2 + 7) or after 6 courses, ie, a subsequent COAP (cyclophosphamide, vincristine, cytarabine, and prednisolone), DAT 2 + 5, and COAP. The relapse risk (81% vs 73%), disease-free survival (16% vs 23%), and overall survival at 5 years (23% vs 22%) did not differ between the 3-course or 6-course arms. In addition to a treatment duration randomization, 362 patients were randomized to receive 12-month maintenance treatment with low-dose interferon, but no benefit was seen with respect to relapse risk, disease-free survival, or overall survival.

Enhancement of cALL immunogenicity by co-culture with a CD154 expressing 293 cell line.

Pre-B cell acute lymphoblastic leukaemia (cALL) commonly occurs in young patients and although successful conventional therapies are available (such as cytotoxic drugs and bone marrow transplantation) for a proportion of patients (approximately 30%) these are ultimately unsuccessful. Recurrence of disease is a result of the failure of the immune system to recognize these abnormal cells and down-regulation of crucial molecules required for cognate CD4(+) T cell recognition has been postulated as a means of immune escape. In this study we show that an embryonic kidney cell line (293 cells) transfected with CD154 (40 L.1) are capable of not only maintaining the viability of primary ALL cells in culture but can also up-regulate the expression of a number of crucial molecules involved in antigen recognition. We show that 40 L.1 cell stimulation of primary ALL cell cultures can not only enhance the allogeneic and autologous MLR response to such cells but will also induce CTL effectors which are capable of lysing wild-type autologous ALL cells. It is therefore conceivable that such an approach could be used to generate an active anti-tumour response in patients, following conventional therapy, reducing the incidence of recurrence.

An analysis of the effect of chronic GvHD on relapse and survival following allogeneic PBSC transplantation.

BACKGROUND: PBSC are increasingly being used as the source of stem cells in allogeneic transplantation. An increased incidence of chronic GvHD has been suggested following unmanipulated allogeneic PBSC transplantation (PBSCT), however, how this affects overall survival is not yet clear. Our aim was to study the impact of chronic GvHD on survival and relapse following allogeneic PBSCT. METHODS: We have analyzed data from 73 patients undergoing HLA-matched allogeneic PBSCT. GvHD prophylaxis was with CYA and MTX in 97% of patients. We have studied the incidence of chronic GvHD and its affect on relapse and survival in these patients. All patients were at least 100 days post-transplant at the time of analysis. RESULTS: Seventy-three patients were evaluable for analysis of chronic GvHD. The overall incidence of chronic GvHD was 55% (limited in 18% and extensive in 37%). Overall median survival was 991 days, with a 4 year survival rate of 48%. Twelve patients relapsed. Patients with chronic GvHD had a significantly lower incidence of disease relapse (p = 0.005) with a relapse probability of 8% at 3 years, compared with 40% in patients with no chronic GvHD. In addition, the extent of chronic GvHD had a marked effect on survival, patients with limited chronic GvHD had a 4 year survival rate of 83%, compared with 45% in patients with extensive chronic GvHD and 38% in patients with no chronic GvHD. This difference was primarily due to the low incidence of relapse and low mortality seen in patients with limited chronic GvHD. DISCUSSION: The presence and extent of chronic GvHD is an important predictor of outcome following allogeneic PBSCT, in that patients who developed either limited or extensive chronic GvHD had a low risk of disease relapse.

Interleukin-4 and tumour necrosis factor-alpha produce non isotype specific partial differentiation of peripheral blood B-cells in myeloma.

In a previous study, culture of peripheral blood mononuclear cells (PBMC) from myeloma patients with interleukin(IL)-4 and tumour necrosis factor(TNF)-alpha resulted in the appearance of clonal plasma cells, thus suggesting the presence of circulating myeloma cell precursors in the peripheral blood. Using the same cytokine combination, we cultured PBMC and purified peripheral blood B-cells from myeloma patients. In nearly all cases, partial differentiation of B-cells occurred but, similarly to results for normal controls, both kappa and lambda light chain (L.C.) cytoplasmic positive lymphoid and lymphoplasmacytoid cells were detected rather than clonal plasma cells. These results suggest that IL-4 and TNF-alpha cause partial differentiation of residual normal polyclonal B-cells rather than of circulating myeloma cell precursors in the peripheral blood of myeloma patients.

Allogeneic peripheral blood stem cell transplantation for haematological malignancies--an analysis of kinetics of engraftment and GVHD risk.

We have carried out an analysis of 44 patients undergoing allogeneic PBSC transplants from fully HLA-matched related donors with particular emphasis on engraftment kinetics and the incidence and severity of GVHD. The recipients had a median age of 37 years (range 5-56 years), 16 patients had standard-risk disease and 28 had poor-risk disease. GVHD prophylaxis was with cyclosporin A and methotrexate (n = 41), cyclosporin A alone (n = 2) or cyclosporin A and methyl-prednisolone (n = 1). Stem cells were mobilised using G-CSF, collecting a median of 5.75 x 10(6) CD34+ cells/kg recipient weight (range 0.94-35 x 10(6) CD34+ cells/kg). Engraftment times to a neutrophil count > 0.5 x 10(9)/1 and platelets > 20 x 10(9)/1 were achieved at a median of day +14 (range 10-25) and day +14 (range 9-130) respectively. Patients receiving > or = 4 x 10(6) CD34+ cells/kg had significantly accelerated neutrophil and platelet engraftment and this number of CD34+ cells would appear to be a prerequisite for maximum engraftment using PBSC. Acute GVHD occurred in 25 of 43 evaluable patients although in only 12 was this clinically significant (grades II-IV). Chronic GVHD has occurred in 17 out of 36 evaluable patients, there was no significant difference between the standard- and poor-risk groups in incidence of either acute or chronic GVHD. In conclusion, these results confirm the feasibility of using PBSC for allogeneic transplantation without evidence for increased risk of either acute or chronic GVHD and provide further evidence supporting the potential of PBSC to replace bone marrow as the major source of haemopoietic cells for allogeneic transplantation.

Microangiopathic Hemolytic Anemia Associated with the use of both Cyclosporin A and FK506 (Tacrolimus) in the Same Patient.

Microangiopathic hemolytic anemia (MAHA) is a well recognized complication of cyclosporin A (CyA) immunosupressive therapy post bone marrow and solid organ transplant but has rarely been reported in association with FK506 (tacrolimus), we describe a unique patient in whom both CyA and FK506 appear to have caused microangiopathy following an allogeneic peripheral stem cell transplant.

G-CSF after peripheral blood stem cell transplantation in lymphoma patients significantly accelerated neutrophil recovery and shortened time in hospital: results of a randomized BNLI trial.

We have undertaken a prospective randomized study in 90 patients with relapsed or resistant lymphomas to assess the value of G-CSF (lenograstim) in the acceleration of myeloid recovery after peripheral blood stem cell transplantation (PBSCT). A common regimen of cyclophosphamide 1.5 g/m2 on day 1 and lenograstim 263 microg s.c. on days 2-10 with two aphereses on days 10 and 11 was used for stem cell mobilization. 77% of patients achieved an adequate PBSC collection in two harvests (> 2 x 10(8) MNC/kg or > 2 x 10(6) CD34+ cells/kg). 65 patients went on to receive high-dose BEAM chemotherapy and engraftment data was available for 62. 34 patients had been randomized to receive lenograstim 263 microg/d s.c. and 28 to no growth factor. The median time to ANC > 0.5 x 10(9)/l was 9 d in the lenograstim arm versus 12.5 d in the no-lenograstim arm (P=0.0001). This was associated with a median duration of time in hospital post PBSCT of 13 d in the lenograstim arm versus 15.5 d in the no-lenograstim arm (P=0.0002). Median days to platelet independence, platelet transfusions, incidence of infection and red cell transfusion were the same in both arms. These data indicate that lenograstim significantly accelerated myeloid recovery after PBSCT and shortened the duration of hospital stay.

Fatal bone marrow embolism in a patient with sickle cell beta + thalassaemia.

Sickle cell beta + thalassaemia is regarded as the mildest of the sickle cell haemoglobinopathy syndromes with a benign natural course. In contrast to sickle cell disease, severe life threatening complications are not usually associated with this genotype. A case of a 30 year old West Indian man who, previously asymptomatic for 10 years, sustained a fatal pulmonary bone marrow embolism, is reported. This case report illustrates that serious, even fatal, complications may occur in patients with this 'benign' condition and bone marrow embolism should be included in the differential diagnosis of acute crisis in these patients.

Successful use of topical retinoic acid in severe dry eye due to chronic graft-versus-host disease.

Topical retinoic acid has proved to be of variable benefit in a number of dry eye disorders of disparate aetiology, in which squamous metaplasia with keratinization of ocular epithelium is present. Its exact role in patients with dry eye however remains in dispute. We describe a case of severe dry eye due to chronic graft-versus-host disease, which was refractory to conventional therapy but which responded remarkably to topical retinoic acid with reversal of conjunctival keratinization and marked resolution of symptoms.

Osteoporosis and vertebral collapse following low-dose, low molecular weight heparin therapy in a young patient.

Osteoporosis is a well-recognized complication of long-term heparin (unfractionated) therapy. It has been suggested that the low molecular weight heparins (LMWH) have a lower potential to cause osteopenia than the unfractionated preparations. We report a case of a 29-year-old woman who developed osteoporosis and pathological fracture of lumbar vertebrae following treatment with low dose LMWH (dalteparin) for 3 months. The aim of this report is to alert clinicians to the potential risk of this complication with the prolonged use of LMWH preparations--an increasingly popular choice for long-term anticoagulation.

Evaluation of the Pall RC100 leucocyte removal filters and the effect of air introduction on its performance.

We evaluated the efficiency of Pall RC100 leucocyte removal filters in processing two units of concentrated red cells for transfusion. Our results show that the filter is very effective in processing the first unit. However, 32% (5/16) of the filters evaluated allowed a leak of substantial numbers of white cells towards the end of the transfusion of the second unit of blood. Air introduction experiments showed that as little as 2 ml of air in the filter may cause major deterioration of the filter function.

Does oxypentifylline reduce the intensity of acute graft-versus-host disease following allogeneic bone marrow transplantation?

From 1988 to 1993, a retrospective analysis of the severity of acute graft-versus-host disease (GVHD) was carried out in two groups of patients undergoing allogeneic bone marrow transplantation for various haematological disorders. One group of 23 patients received oxypentifylline in a dose of 400 mg, 6 hourly, orally, day -10 to day +35 in addition to other standard management received by the control group of 20 patients. Acute GVHD was scored in each patient according to the Seattle criteria. Patients receiving oxypentifylline showed a lower GVHD score in each organ-specific area and this reached significance with the total GVHD score. A significantly smaller proportion of patients in the treatment group developed acute GVHD of grade 2 or more. It is concluded that oxypentifylline used in the present dosage reduces the intensity of acute GVHD following allogeneic bone marrow transplantation and is also well tolerated by the patient.

Thrombocytopenia following autologous bone marrow transplantation: evidence for autoimmune aetiology and B cell clonal involvement.

Thrombocytopenia outlasting anaemia and neutropenia is a well recognised sequel of autologous bone marrow transplantation (BMT) but the pathogenesis remains unclear. Autoimmune destruction of platelets has been suggested as a possible mechanism. We studied 5 patients who had undergone autologous BMT and were found to have persistent thrombocytopenia (< 150 x 10(9)/l) 6 months from transplantation with a normal haemoglobin level and granulocyte count. Apart from a mild reduction in the megakaryocyte numbers in one case, no other quantitative or qualitative defects of the megakaryocyte lineage were present to explain the peripheral thrombocytopenia. Two cases had positive anti-platelet autoantibodies. Immunoglobulin heavy chain gene rearrangement studies of peripheral blood and bone marrow mononuclear cells using the polymerase chain reaction showed evidence of clonal rearrangement in one of the two cases with positive anti-platelet autoantibodies. Our results support the previous reports that anti-platelet antibody-mediated destruction of platelets may play a role in the pathogenesis of post-autologous BMT thrombocytopenia. Furthermore, the demonstration of a clonal B cell expansion in one of the cases with anti-platelet antibodies suggests an aetiological link between clonal B cells, autoantibody production and thrombocytopenia.

Successful pregnancies in women following single autotransplant for acute myeloid leukemia with a chemotherapy ablation protocol.

Of 30 women surviving a minimum of 18 months following treatment for AML with a high-dose chemotherapy regimen with autologous bone marrow transplantation (ABMT), 24 were premenopausal at the time of transplantation. All were given a detailed questionnaire concerning menstruation, menopausal symptoms and pregnancy; 22 responded. Of these 22, 10 had received a single transplant procedure and 12 a double transplant procedure. In the 10 recipients of a single transplant, 4 women (age range 32-50 years) developed ovarian failure and 6 (age range 21-32 years) resumed spontaneous cyclical menstruation. Five of the 6 menstruating women became pregnant between 4 and 40 months following ABMT. Three pregnancies went to term and each resulted in the delivery of a full-term apparently normal infant. Of the 12 women who received a double ABMT (age range 32-47 years), 11 developed clinical and/or biochemical evidence of ovarian failure. The median age in the latter group was 35 years, however, compared with 28 years in the single ABMT group. These data show that it is possible to give a single very high-dose course of chemotherapy in younger patients without compromising fertility.

Identification and treatment of anaemia in older patients.

Anaemia in elderly patients should never be regarded as a normal physiological response to aging. Underlying causes must be investigated and treated in a similar manner to that used in younger adults. In addition to a thorough history and physical examination, basic investigations such as red cell indices and morphology, reticulocyte count, haematinic assays and occasionally bone marrow examination, will detect the underlying pathology in most cases. Anaemia may be classified, according to red blood cell mean corpuscular volume, into microcytic, macrocytic and normocytic types. Anaemia with an absolute reticulocytosis is due either to acute blood loss or haemolysis. Other anaemias, more frequently encountered in elderly patients, are hypoproliferative, and reflect depressed marrow production or impaired erythroid maturation. Examples include anaemia of chronic disease and iron deficiency and, less commonly, megaloblastic anaemia and anaemia due to primary bone marrow failure. The treatment of anaemia should aim to correct the underlying cause of the disorder and/or to improve the quality of the blood, e.g. by haematinic replacement therapy. Recombinant human erythropoietin has revolutionised the treatment of anaemia associated with chronic renal failure, while its role in other anaemias is currently under investigation. Regular blood transfusion may be required for some elderly patients with chronic anaemia. However, the attendant risks of this procedure, such as iron overload and viral hepatitis transmission, must be considered.

Pulmonary complications of bone marrow transplantation: the impact of variations in total body irradiation parameters.

In this article we report and discuss the pulmonary complications in patients who received a single exposure total body irradiation (TBI) to a total dose of 7.5 Gy at a dose rate of 0.15 Gy/min, a TBI regimen which has the advantage of being given in a single, relatively short exposure, with an active treatment time of less than 1 hour. This forms a part of the bone marrow transplantation programme for the management of certain haematological malignancies at the Leicester Regional Centre. Between July 1986 and October 1990, we treated 31 patients with such a regimen. Full respiratory function tests (RFT) were carried out, prior to TBI, in the majority of patients. After a mean follow-up period of 34 months, 13 patients were alive; full RFT were repeated in all of them. Of the total of 31 patients, only one patient died, from late non-specific pneumonitis; in this case, high dose busulphan was added to conventional cyclophosphamide and TBI. Another patient died as a direct result of cytomegalovirus pneumonia. Comparison of pre- and post-TBI RFT showed no resultant obstructive, restrictive or transfer factor defects. In the three patients who did not have pre-TBI RFT, post-TBI RFT did not reveal any significant change from expected values for age, sex and height. Several major centres have reported their experience using various combinations of different total doses, dose rates and fractionations. Having compared our results with theirs, we conclude that, following this relatively short and convenient single exposure TBI, there is no evidence of increased acute or chronic pulmonary toxicity.

Flow cytometric quantitation of sequence-specific mRNA in hemopoietic cell suspensions by primer-induced in situ (PRINS) fluorescent nucleotide labeling.

We modified a technique of RNA detection based upon primed in situ labeling of RNA (PRINS) to quantitate poly(A) and histone mRNA in HL60 cell suspensions by flow cytometry. The direct PRINS technique was used to label poly(A) mRNA, based upon the incorporation of fluoresceinated nucleotides into cDNA chains generated by reverse transcription from the site of oligo(dT) primer-specific annealing on the poly(A) template. By this method, poly(A) fluorescent signals of up to 20-fold above background were obtained when primer concentrations were saturating. Nonspecific binding of the fluorochrome-conjugated nucleotide proved to be the greatest source of background fluorescence in cell suspensions. This was considerably reduced by preincubation of the cells in lysine and an excess of cold nucleotides prior to the PRINS reaction. The application of indirectly fluoresceinated antibodies to the FITC moiety of the conjugated nucleotide (indirect PRINS) enhanced detection of a fluorescent signal generated from cells by the direct PRINS technique after stringent histone priming. Cell cycle analysis, based upon DNA histograms obtained after PRINS labeling, showed the histone fluorescence to be primarily in S-phase cells of an exponentially growing population. Confocal laser scanning microscopy revealed good retention of cellular architecture, with cytoplasmic localization of poly(A) and histone mRNA fluorescence and heterogeneous mRNA expression among cells. Our modified PRINS labeling of RNA represents a useful advance in the precise quantitation of RNA species in cell suspension by flow cytometry.