S. J. Proctor

The relationship between bcl‐2 expression and response to chemotherapy in acute leukaemia

Summary. Immunocytochemistry was used to assess bcl‐2 expression in blasts obtained from the bone marrow of 28 patients with acute lymphoblastic leukaemia (ALL) (16 children and six adults at presentation and three children and three adults on relapse) and 20 with acute myeloid leukaemia (AML) (19 adults and one child, 13 with de novo AML, 11 at presentation and two on relapse, and seven secondary to myelodysplasia or chronic myeloid leukaemia). Slides were examined both for the percentage of positive cells and for the intensity of staining using a five‐point scale. There was a statistically significant increase in both the percentage of positive cells seen (P < 0.002) and the intensity of staining (P < 0.01) between samples obtained at relapse and those at presentation in ALL. There was a significantly greater intensity of staining in cells from patients with ALL (P < 0.05) and AML (P < 0.05) who failed to achieve remission after chemotherapy than in those who responded. The intensity of staining in cases of secondary AML was lower than that in de novo disease (P < 0.01). These results suggest that expression of bcl‐2 may be an important prognostic feature in both de novo AML and in ALL, but not in secondary AML.

Non-Hodgkin's Lymphoma and Hepatitis C Virus Infection

The hepatitis C virus (HCV) is a recently described and important cause of acute and chronic liver disease. A hallmark of HCV is its propensity to become chronic, some patients with chronic HCV progressing to cirrhosis and hepatocellular carcinoma (HCC). HCV is also lymphotrophic and we report 2 patients with HCV cirrhosis who developed non-Hodgkins lymphoma (NHL). These cases raise the possibility that chronic HCV infection of lymphocytes plays an aetiological role in this malignancy. However screening of a further 63 consecutive patients over the age of 50 years with NHL for HCV antibody by second generation enzyme linked immunoassay (ELISA) failed to identify any patients with evidence of HCV infection. This suggests that HCV is an uncommon contributory factor for the development of non-Hodgkins lymphoma in the United Kingdom.

Somatic mitochondrial DNA mutations in adult-onset leukaemia

Mitochondrial genome instability has recently been demonstrated in a wide variety of human tumours and is implicated in the development of the myelodysplastic syndromes, a heterogeneous group of haematological disorders with an increased risk of malignant transformation. We therefore investigated the incidence of somatic mitochondrial DNA (mtDNA) mutations in patients with adult-onset leukaemia. We sequenced the entire mitochondrial genome from both normal tissue (buccal epithelial cells) and the leukaemia from 24 patients with adult-onset leukaemia. Somatic mtDNA mutation was present in nine individuals (≈40%) and in each case the tumour genome differed from the normal genome sequence by a single sequence change. Using PCR-RFLP analysis and real-time PCR, we have studied in detail the mutation present in one patient with acute lymphatic leukaemia, demonstrating that the mutation is associated specifically with the leukaemia.

Natural killer cell activity in childhood acute lymphoblastic leukaemia in remission

Summary. Fifteen children with acute lymphoblastic leukaemia (ALL) in remission receiving maintenance chemotherapy and 12 ALL patients off treatment and in remission were tested for natural killer (NK) cell activity in vitro. Compared with a control population the children with ALL receiving maintenance chemotherapy had low levels of NK cell activity. This effect was not due to a specific reduction in NK cell numbers since proportions of mononuclear cells detected by the monoclonal antibodies HNK‐1 (Leu‐7) and Leu‐11a were normal. Furthermore NK cell activity in patients could only be partially increased by pre‐incubation of effector cells with interferon (αIFN). These studies confirm the lack of NK cell activity in children with ALL and show that this phenomenon is directly related to functional NK cell impairment. Our study has further shown that this effect is transient since ALL patients off treatment and in remission showed normal levels and augmentation of NK cell activity.

Corticosteroid resistance is increased in lymphoblasts from adults compared with children: preliminary results of in vitro drug sensitivity study in adults with acute lymphoblastic leukaemia

Summary The prognosis of acute lymphoblastic leukaemia (ALL) in adults is poor compared with children in terms of complete remission (CR) and leukaemia‐free survival. In children in vitro resistance of leukaemic cells to various cytotoxic agents is an independent poor prognostic marker, but the relevance of in vitro drug resistance in adults to poor prognosis has not been described. Lymphoblasts from 16 adults and 32 children with ALL at initial presentation were assayed for in vitro drug sensitivity in a short‐term culture system using the reduction of 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide (MTT) as an indicator of cell viability. The following drugs were tested: prednisolone, daunorubicin, mitozantrone, etoposide, melphalan and 6‐thioguanine. At initial presentation, lymphoblasts from adults demonstrated a significantly higher degree of in vitro resistance to prednisolone than those from children (P < 001). Glucocorticoid resistance may be a fundamental difference between adult and childhood ALL which may underlie different biological aspects and also explain the difference in prognosis. Lymphoblasts from adults who achieved CR were more sensitive in vitro to prednisolone (P = 007), daunorubicin (P < 005), mitozantrone (P < 001) and melphalan (P < 005) than cells from those who did not. The MTT assay can predict response to induction chemotherapy in adults and therefore discriminate between standard‐ and high‐risk patients. The assay, however, is not suitable for selection of the most effective agent for treatment because of in vitro cross‐resistance of lymphoblasts to various drugs tested.

Bone marrow transplantation for acute myeloblastic leukaemia: an EBMT Leukaemia Working Party prospective analysis from HLA-typing.

Summary. The optimal post‐remission therapy for patients with acute myeloblastic leukaemia remains controversial. Allogeneic bone marrow transplantation, autologous bone marrow transplantation, and consolidation chemotherapy are the major options. In order to evaluate their respective value the European Group for Bone Marrow Transplantation conducted a prospective registration study. Patients with newly diagnosed acute myeloblastic leukaemia were registered at the time of HLA‐typing and intention to treat in case of presence or absence of an HLA‐identical donor was recorded. 27/79 (34%) patients HLA‐typed at diagnosis had an identical donor identified. The estimated survivals at 3 years from HLA‐typing were 44% and 21% among patients with or without HLA‐identical donor, respectively (P= 0·02). 22/26 (85%) patients for whom allogeneic bone marrow transplantation was intended were transplanted but only 15/47 (32%) patients for whom autologous bone marrow transplantation was intended were indeed transplanted (P < 0·001). The survival was 50%. 29% and 17% (P= 0·004) for patients treated with allogeneic bone marrow transplantation, autologous bone marrow transplantation, or chemotherapy, respectively. 40/68 patients HLA‐typed in first complete remission had an HLA‐identical donor. The estimated 3‐year survival among patients typed in first remission with and without HLA‐identical donors was 42% and 35% (n.s.), respectively. This technique of early patient registration illustrates the problems of patient selection during the course of the disease and might be used as a complement to randomized trials when comparing bone marrow transplantation and other treatment options.

Low incidence of myelodysplastic syndrome following transplantation using autologous non-cryopreserved bone marrow.

Between May 1984 and October 1995 we performed 114 autologous stem cell transplants for lymphoma in our centre; 77/114 (68%) were transplanted after primary therapy. The conditioning regimen varied according to diagnosis; 26 patients were conditioned with melphalan and total body irradiation, 66 received melphalan and etoposide and the remainder (50) were conditioned with melphalan alone. The median follow-up is 62 months. Only two new haematological malignancies have occurred, both in patients with Hodgkin’s disease. One patient developed Ph+ chronic myeloid leukaemia 18 months post-transplant. In this case, because of the timing of the haematological disorder, we considered the malignancy to be concurrent with or to have preceded the transplant. A second patient developed acute myeloid leukaemia 20 months post-transplant. She had been treated for Hodgkin’s disease for 10 years and was transplanted in third complete remission. Cytogenetic analysis in this case showed trisomy 11. We believe this to have been an unequivocal second malignancy. Our finding of a 1.1% incidence of secondary haematological malignancy (95% CI 0.02–4.96) from a census population adds weight to the hypothesis that haematological problems post-transplant reflects prior chemotherapy rather than toxicity from the transplant procedure itself.

A comparative study of combination chemotherapy versus marrow transplant in first remission in adult acute lymphoblastic leukaemia

Summary. The results of conventional chemotherapy in adult acute lymphoblastic leukaemia (ALL) have not improved substantially in recent years. The present study is based on a flexible policy of marrow transplantation (allograft and autograft without marrow purging) in first remission compared with a group treated with standard maintenance therapy after a common induction sequence. The actuarial disease free survival (DFS) and actuarial overall survival (OS) at 3 years for autologous marrow grafted patients was 30% and 65% respectively. The allogeneic transplant group had DFS of 30% and OS at 3 years of 38% compared with DFS (12%) and OS (12%) for patients on 6‐mercaptopurine and methotrexate maintenance. The actuarial disease free survival calculations include patients on protocol not entering remission, therefore, giving the worst possible result.

Tumour Necrosis Factor Gene Polymorphisms in Lymphoproliferative Disease

Tumour necrosis factor (TNF) α is involved in the pathogenesis of established lymphoproliferative disease. Serum levels of TNFα and its soluble receptors are above normal values in B-cell chronic lymphocytic leukaemia (B-CLL) and they are valuable prognostic markers in lymphoma patients. The production of TNFα is genetically controlled. Altered synthesis of TNFα has been associated with polymorphisms at the TNF gene cluster (i.e. TNFA, TNFB and LTB). In the present study, we evaluated the prevalence of the known high TNFα- and TNFβ- producing alleles TNF1, TNF2 of the TNFA gene, TNFB1, TNFB2 alleles of the TNFB gene and of the polymorphic alleles TNFd1, d2, d3, d4 and d5 of the microsatellite TNFd in patients with B-CLL, non-Hodgkins lymphoma (NHL) and Hodgkins disease (HD). This study demonstrates that there is no difference in the frequency of the tested TNF alleles between normal controls and cohorts of patients with lymphoproliferative disease. These results indicate that TNF alleles are not genetic predisposing factors in the development of these diseases.

Inhibition of natural killing and antibody-dependent cell-mediated cytotoxicity by the plasma protease inhibitor α2-macroglobulin (α2M) and α2M protease complexes

Over the past few years increasing attention has been given to the relationship between the immune response and proteases. The aim of our present study was to examine the dose-response effect of purified alpha 2-macroglobulin (alpha 2M) with varying degrees of protease (trypsin) saturation on natural killing (NK) and antibody-dependent cell-mediated cytotoxicity (ADCC). The results demonstrated that alpha 2M with 50% trypsin saturation (fast alpha 2M) was more inhibitory in both assays than alpha 2M with no bound protease (slow alpha 2M).