BACKGROUND:Chronic graft-versus-host disease (GVHD) is the leading cause of later illness and death after allogeneic hematopoietic stem-cell transplantation. We hypothesized that the inclusion of antihuman T-lymphocyte immune globulin (ATG) in a myeloablative conditioning regimen for patients with acute leukemia would result in a significant reduction in chronic GVHD 2 years after allogeneic peripheral-blood stem-cell transplantation from an HLA-identical sibling.METHODS:We conducted a prospective, multicenter, open-label, randomized phase 3 study of ATG as part of a conditioning regimen. A total of 168 patients were enrolled at 27 centers. Patients were randomly assigned in a 1:1 ratio to receive ATG or not receive ATG, with stratification according to center and risk of disease.RESULTS:After a median follow-up of 24 months, the cumulative incidence of chronic GVHD was 32.2% (95% confidence interval [CI], 22.1 to 46.7) in the ATG group and 68.7% (95% CI, 58.4 to 80.7) in the non-ATG group (P<0.001). The rate of 2-year relapse-free survival was similar in the ATG group and the non-ATG group (59.4% [95% CI, 47.8 to 69.2] and 64.6% [95% CI, 50.9 to 75.3], respectively; P=0.21), as was the rate of overall survival (74.1% [95% CI, 62.7 to 82.5] and 77.9% [95% CI, 66.1 to 86.1], respectively; P=0.46). There were no significant between-group differences in the rates of relapse, infectious complications, acute GVHD, or adverse events. The rate of a composite end point of chronic GVHD-free and relapse-free survival at 2 years was significantly higher in the ATG group than in the non-ATG group (36.6% vs. 16.8%, P=0.005).CONCLUSIONS:The inclusion of ATG resulted in a significantly lower rate of chronic GVHD after allogeneic transplantation than the rate without ATG. The survival rate was similar in the two groups, but the rate of a composite end point of chronic GVHD-free survival and relapse-free survival was higher with ATG. (Funded by the Neovii Biotech and the European Society for Blood and Marrow Transplantation; ClinicalTrials.gov number, NCT00678275.).

BACKGROUND AND OBJECTIVES In acute leukemias, chromosomal translocations involving the 11q23 band are frequently, but not invariably, associated with MLL gene rearrangement and their finding is associated with a poor prognosis. We observed two new translocations with a breakpoint in the 11q23 region at standard cytogenetic analysis: a previously undescribed t(3;11)(q21;q23) in a 70-year old woman with a fulminating form of AML-M1 and a new translocation t(6;11)(q15;q23) in a 61-year old man with an atypical chronic myelogenous leukemia. In these two patients, involvement of the MLL gene was analyzed by molecular cytogenetic techniques which also allowed a more precise mapping of the breakpoints. DESIGN AND METHODS The MLL gene was analyzed by Southern blot and by fluorescent in situ hybridization (FISH) with a double-color MLL probe. A panel of 11q, 3q and 6q cosmid/YAC probes mapping around the breakpoints was used for breakpoint mapping. RESULTS In both patients, FISH analysis and Southern blot showed that the MLL gene was not rearranged; in patient 1, MLL was retained on the 11q+ derivative, whereas in patient 2 it moved to the 6q- chromosome. In the t(3;11) we localized the chromosome 11 breakpoint at 11q23.3, in a region flanked by CP-939H3 and cos1p3, distal to the MLL locus; in the t(6;11) the break occurred at 11q21, in a region flanked by CP-819A5 and CP-829A6, proximal to the MLL locus. INTERPRETATION AND CONCLUSIONS Our cases add two new translocations to the list of chromosomal anomalies involving the long arm of chromosome 11, and show that apparent translocation t(11q23) may involve loci and genes other than MLL. Characterizing the molecular heterogeneity of 11q23 translocations may identify some prognostic significance.

Serum levels of tumour necrosis factor-alpha (TNF-alpha) have been evaluated in the peripheral blood of 91 patients with B-cell chronic lymphocytic leukaemia (B-CLL), and have been correlated with the clinical stage (according to Rai's staging system) and relevant haematological and immunological data. Increased values were detected, compared to 36 normal age-matched controls (36 pg/ml +/- 5 versus 0.11 pg/ml +/- 0.08; P < 0.05). An increase of TNF-alpha serum levels was observed in all stages including stage 0, with a progressive increase in relation to the stage of the disease. A significant relationship between serum TNF-alpha levels and the number of circulating monocytes (P < 0.002) and an inverse correlation with the level of the haemoglobin (P < 0.001) was established, as defined by the Pearson's correlation test. In contrast, no correlation was observed between TNF-alpha serum levels and the other parameters taken into account, including the white blood cell and platelet counts, the absolute number of peripheral blood (PB) lymphocytes, CD5+ B lymphocytes, CD57+ lymphocytes, serum levels of lactic dehydrogenase, total serum immunoglobulins and the serum levels of IgG, IgA and IgM. These data suggest that, in addition to the B-CLL neoplastic cells, the PB monocytes may be involved in the release of TNF-alpha.

To extent our knowledge on the cytokines possibly involved in the pathophysiology of B-cell chronic lymphocytic leukemia (B-CLL), the mRNA expression of a panel of 10 cytokines was investigated on purified B-CLL cells using a reverse-transcriptase polymerase chain reaction method. Whereas negative RT-PCR signals were recorded for interleukin-1 alpha (IL-1 alpha), IL-2, IL-3, IL-4, IL-5, IL-7, tumor necrosis factor beta (TNF beta), and granulocyte-macrophage colony-stimulating factor, we detected the expression of IL-1 beta, IL-6 and TNF alpha. Furthermore, the constitutive expression of IL-8 mRNA was observed in all 17 B-CLL samples analyzed. mRNA expression was associated with the capacity of the leukemic cells to release IL-8 both constitutively (4.6 +/- 8.1 SD ng/mL) and, to a further extent, after stimulation (14.5 +/- 19.4 ng/mL). The circulating levels of IL-8 were also evaluated in 12 untreated B-CLL sera samples and the overall mean level was significantly higher (P <.01) than in normal sera. In addition, supernatants of purified B-CLL cells cultured in the presence of 12-O- tetradecanoylphorbol-13-acetate showed chemotactic activity towards neutrophils; this activity was neutralized in the presence of an anti- IL-8 antiserum. The mRNA for IL-8 was absent in five B-cell preparations from hairy cell leukemia cases and in four B-cell lines. Normal tonsil CD5+ B cells showed a low expression of IL-8 mRNA only in two of the nine preparations tested and the overall quantity of IL-8 released by these cells after 3 days' incubation was significantly lower compared with that released by B-CLL cells (0.4 +/- 0.3 and 1.6 +/- 0.9 ng/mL under basal and stimulated conditions, respectively). These findings point to an involvement of a member of the proinflammatory chemokine supergene family in human CD5+ B lymphocytes. The different IL-8 behavior observed between B-CLL cells and their normal counterpart is likely to reflect an activation state of the leukemic population.

We hereby here the successful use of a combined therapeutic approach with alpha 2b Interferon (IFN) and Chlorambucil in the management of previously treated B-cell chronic lymphocytic leukemia (B-CLL). The two patients under study had been managed with repeated and frequent cycles of Chlorambucil. Rather than with more toxic multidrug regimens, the hematological picture was controlled with 3 MU of IFN every three-five days and 5 mg of Chlorambucil every two-five days. Using such an approach, which is feasible on an out-patients basis and devoid of significant side effects, a consistent control of the WBC count was achieved for 32 and 16 months, respectively.

The possibility of evaluating the susceptibility of B-cell chronic lymphocytic leukemia (B-CLL) cells to chemotherapeutic agents pre-clinically, was assessed using the MTT colorimetric assay. The results so far obtained suggest that this rapid 3-4 days' test is accurate and reliable and may allow the physician to recognize individual B-CLL cases in which the neoplastic clone is resistant to Chlorambucil. Furthermore, the high efficacy of Fludarabine clearly emerges from these in vitro analyses. The possibility of employing Chlorambucil in combination with alpha Interferon is suggested and the relevance of using this simple test for better drug selection in B-CLL patients is discussed.