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Objectives: Mutations in the C-terminal region of the nucleophosmin (NPM1) gene occur in approximately 60% of acute myeloid leukemia (AML) cases with normal karyotype and represent the most common genetic lesion presently known in this disease. Because of their frequency and favorable impact on prognostic outcome, screening for this aberration is currently recommended in routine diagnostic characterization of AML. Several techniques enabling to detect NPM1 mutation have been reported, but all require sophisticated equipment, which represent an obstacle particularly in countries with limited resources., Methods: We designed an RT-PCR strategy to amplify NPM1 exon 12 followed by electrophoresis and fragment visualization on polyacrylamide gels to discriminate a 4-5 bp size difference resulting from mutations in this gene. A hemi-nested method was designed to increase sensitivity for the study of minimal residual disease (MRD)., Results: The assay enabled specific detection of NPM1 mutations in 12/36 patients. A 10(-2) sensitivity level was obtained using one amplification round, while the hemi-nested PCR approach yielded a 10(-5) sensitivity level, therefore proving useful to assess MRD in patients carrying the mutation. The results were independently validated in 24 AML cases by sequencing analysis., Conclusions: This simple and low-cost assay may integrate diagnostic work-up of AML and could be used for assessment of response to therapy in patients with NPM1 mutations.

Objectives: An aberrant pattern of expression of L-selectin and intercellular adhesion molecule 1 (ICAM1) may characterise CD34+ blast cells in myelodysplastic syndromes (MDS) and secondary acute myeloid leukaemia (sAML)., Methods: In a three-colour flow cytometric assay, we evaluated the expression of L-selectin and ICAM1 on CD34+ blast cells from the bone marrow (BM) of 66 MDS patients; for the purpose of comparison CD34+ blast cells of 18 sAML and CD34+ stem cells of 17 normal donors were also analysed., Results: The ratio of L-selectin/ICAM1 expression was identified as a parameter correlated with the percentage of BM blast infiltration and the time to leukaemic progression among MDS patients. In fact, the values of L-selectin/ICAM1 ratio were inversely correlated with the BM blast infiltration (r = -0.34, P = 0.004). Furthermore, MDS patients with a baseline ratio <1 had a higher leukaemic progression rate (41% vs. 19%, P = 0.008); the actuarial risk of disease progression for this subgroup of MDS patients was also higher (64% vs. 11% at 2 yr, P = 0.002). Furthermore, in two patients a decrease of the ratio was observed when overt leukaemic transformation occurred; conversely, restoration of a normal ratio was observed in two patients after a chemotherapy-induced remission., Conclusion: (i) L-selectin is defective in the stem cell compartment of MDS and sAML, whereas ICAM1 is overexpressed; (ii) the ratio of their expression has a prognostic role; and (iii) a ratio <1 significantly predicts progression to overt leukaemia in MDS patients.

Background: Splenic marginal zone lymphoma (SMZL) is an infrequent B-cell neoplasm that pursues an indolent course. Signs and symptoms, mostly related to hypersplenism, are successfully managed by splenectomy. However, the therapy of patients who are not fit for a surgical procedure or who relapse after splenectomy, is still an unsettled issue., Patients and Methods: We report a phase-II study on 16 patients with SMZL, three therapy naïve and 13 pretreated, all showing systemic symptoms or progressive worsening of peripheral cytopenia, who were treated with pentostatin at a dose of 4 mg/m2 every other week for 6-10 wk. In relapsed patients, the median interval between diagnosis and treatment was 26 month (range: 8-49)., Results: Overall, 68% of the patients showed a clinical response. Two out three patients, who received pentostatin as first line therapy, attained a complete response (CR). One CR and seven minor or good haematological responses were recorded in relapsed patients. Treatment toxicity, mostly haematological, proved manageable. With a median follow-up of 35 month the median overall survival (OS) is 40 month and the median progression free survival (PFS) is 18 month., Conclusion: Our data show that pentostatin administered every other week has a good degree of activity in the treatment of SMZL and suggest that this schedule could be considered a possible therapeutic option for patients who are not fit for splenectomy or have relapsed., (Copyright Blackwell Munksgaard 2005.)

Reactivation of hepatitis B virus infection in subjects receiving cytotoxic treatment for heamatological malignancies occurs in 21-53% of chronic HBsAg carriers and in an unknown number of HBsAg negative subjects harbouring occult HBV infection. Immunotherapy with alemtuzumab, a humanized monoclonal antibody against CD52 epitopes on lymphocytes cells produces deep immunosuppression. We describe two subjects with chronic lymphocytic leukaemia and occult HBV infection who developed a virological and biochemical flare of hepatitis B following immunotherapy with alemtuzumab. One of them developed full blown hepatitis with seroreversion from anti-HBs to HBsAg after four weeks of alemtuzumab therapy. Lamivudine (100 mg die) achieved a complete clinical recovery and HBV-DNA clearance from blood within 8 weeks. The second patient (HBsAg and HBV-DNA seronegative, anti-HBs and anti-HBc positive before treatment) was kept under prophylaxis with lamivudine up to three months after alemtuzumab. Two months after withdrawal of lamivudine, clinical and laboratory features of acute hepatitis B developed. Lamivudine therapy was restarted and a prompt recovery was obtained with HBsAg and HBV-DNA clearance.