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Introduction. Acute myeloid leukemia (AML) is a heterogeneous disease with various chromosomal aberrations. The karyotype at diagnosis provides important prognostic information that influences therapy and outcome of this disease. However, using conventional chromosome banding techniques alone, karyotype abnormalities are detected in only half of all AML cases. Aims.We sought to identify novel genomic regions of interest in normal karyotype AML and to identify novel candidate regions and disease-related genes in patients with complex karyotypes using genome-wide high resolution SNP-array. Patients and Methods. Samples from 57 AML patients with FAB-M0, M1, M2, M3, M4, M5, miscellaneous cytogenetic abnormalities and normal karyotype were examined. Genomic DNA was isolated from mononuclear AML cells and applied to GeneChip® Human Mapping 250K NspI and Genome-Wide Human SNP 6.0 array microarrays (Affymetrix, Santa Clara, CA) following the manufacturer’s instructions. Copy number aberrations were scored using the Hidden Markov Model and the segmentation approach available within the Partek software package as well as by visual inspection. All aberrations were calculated with respect to a set of 270 Hapmap normal individuals and a set of samples obtained from acute leukaemia cases in remission in order to reduce the noise of raw copy number data. When available, in order to exclude inherited copy number variants a comparison to paired constitutional DNA and to paired remission DNA was performed. Fluorescence in situ hybridization, quantitative PCR and nucleotide sequencing were used to confirm genomic alterations. Results. A wide spectrum of different genetic lesions (gains/losses) involving complete chromosome arms (del 16q, i(13q10), del 3p, del 7p, monosomy 9) or submicroscopic genomic intervals were identified in a substantial proportion of cases (55%) without differences in the frequency of losses or gains. Focal genetic alterations were detected at the breakpoints of previously cytogenetically identified chromosomal translocations, such as t(2;3)(p22-23)(q26-27) and t(1;11)(p32;q23). Hemizygous deletions were identified at 2q33.3-q34 involving ERBB4 (verb-a erythroblastic leukemia viral oncogene homolog 4 avian), at 9p21.3- p21.2 (CDKN2A-2B), at 12p13 (ETV6), at 17q12 (NF1) and 21q21.2 (RUNX1). Most frequent alterations affected the oncogene MYC at 8q24.13 - q24.21 (4.33 Mb), the ABC transporters genes ABCA8, ABCA9, ABCA6, ABCA5, ABCA10 at 17q24, PTPRM at 18p11.31-p11.23 and ERG at 21q22. Other recurring genetic lesions were uncommon and were identified only in single cases. Some lesions affected regions with a single gene, such as: ETAA1, FIGN, STK32B, PRAGMIN, PCM1, GLIS3, MRGPRX1, SESN3, BCL2L14 or lacking annotated genes. Marked differences in the combination of copy number anomalies were identified across the different genetic subtypes of AML. Patients with normal karyotype showed no relevant genetic alterations. Conclusions. These data demonstrated that, in contrast to adult acute lymphoblastic leukaemia (ALL), AML is characterized by relatively few recurring copy number alterations, and that spectrum of genetic anomalies is significantly associated with AML disease subtype. Supported by: European LeukemiaNet, AIL, AIRC, FIRB 2006, Fondazione del Monte di Bologna e Ravenna, PIO project 2007, Strategico di Ateneo.

We performed a survey of the European Cooperative Group for Blood and Marrow Transplantation to analyze the outcome of 625 acute promyelocytic leukemia (APL) patients transplanted with auto- or allogeneic-hematopoietic stem cell transplantation (autoHSCT, alloHSCT) after 1993, in first (CR1) or in second complete remission (CR2). Leukemia-free survival (LFS) at 5 years in CR1 was 69% for 149 patients autografted and 68% for 144 patients allografted, whereas in CR2, LFS was 51% in 195 autoHSCT and 59% in 137 alloHSCT recipients, respectively. In the group of autoHSCT for CR1 (n=149), higher relapse incidence (RI) was associated with shorter time from diagnosis to transplant (7.6 months); transplant-related mortality (TRM) was increased in older patients (47 years), whereas for CR2, longer time from diagnosis to transplant (18 months) was associated with increased LFS and decreased RI. In the alloHSCT group for CR1 (n=144), age (33 years) was associated with increased LFS and decreased TRM and for CR2 (n=137), the use of mobilized peripheral blood stem cells was associated with decreased TRM. Female recipient, a female donor to male recipient and transplants performed before 1997 were associated with decreased RI. In conclusion, HSCT still appears to have a role in APL, especially for patients in CR2.

In spite of the very high cure rate (70%-80%) achieved in APL with combinatorial all-trans retinoic acid (ATRA) and anthracycline-based chemotherapy regimens, a number of issues are still open for investigation in front-line therapy of this disease. These include, among others, improvements in early death rate, the role of arsenic trioxide (ATO) and maintenance treatment, and, finally, optimization of molecular monitoring to better identify patients at increased risk of relapse. The current consensus on the most appropriate induction therapy consists of the concomitant administration of ATRA and anthracycline-based chemotherapy. Although the antileukemic benefit provided by the addition of ATRA to consolidation therapy has not been demonstrated in randomized studies, historical comparisons of consecutive studies carried out by Spanish and Italian cooperative groups suggest that the combination of ATRA and chemotherapy for consolidation may also contribute to improving therapeutic results. While a variety of distinct treatments are being investigated for front-line therapy, most experts agree that a risk-adapted therapy represents the optimal approach, through the use of more intensive therapy in patients with initial hyperleukocytosis. Longitudinal RT-PCR of PML/RARalpha allows sensitive assessment of response to treatment and minimal residual disease (MRD) monitoring in APL. Achievement of negative PCR status or molecular remission at the end of consolidation is now universally accepted and recommended as a therapeutic objective in this disease. On the other hand, persistence of, or conversion to, PCR positive in the marrow during follow-up is associated with impending relapse. Preliminary studies on therapy of molecular relapse indicate a survival advantage as compared to administering salvage treatment at time of hematologic relapse. The more accurate and reproducible real-time PCR method to detect at quantitative levels the PML/RARalpha hybrid will likely provide better inter-laboratory standardization and trial results comparison in the near future.

Il presente studio si inserisce in una ricerca pluriennale che stiamo conducendo sul Golfo di Trieste[1], un comparto ambientale che per la sua particolare morfologia si è dimostrato sensibilmente vulnerabile. Si tratta infatti di un'area ristretta dell'Alto Adriatico di scarsa profondità (10-20 m) e ricambio idrico modesto (ad eccezione di giornate di elevata ventosità con venti di bora ENE), che riceve gli sversamenti urbani, portuali ed agricoli di Trieste e Monfalcone, nonché contributi antropici fluviali (fiume Isonzo) anche dalla Slovenia. Particolarmente critiche le zone lagunari di Grado e Marano, e del porto di Trieste racchiuso da un sistema di dighe foranee, in quanto corpi acquatici chiusi e statici. Lo studio si è focalizzato sull'area circostante il terminale del diffusore che convoglia offshore le acque trattate dall'impianto di depurazione dei reflui urbani ed industriali della Provincia di Trieste. Ai fini di una migliore comprensione del ruolo che le specie complessanti hanno nel trasporto dei metalli nell'ambiente marino e per studiare la potenziale tossicità di questi ultimi nei confronti degli organismi marini, abbiamo valutato il ruolo degli acidi umici e fulvici, complessanti ubiquitari nell'ambiente marino, rilevanti sia in processi microbici, sia per loro interazione con specie tossiche, metalli, nutrienti ed alogeno-organici. A tal fine si sono effettuati vari campionamenti nel golfo scegliendo punti con caratteristiche spaziali e temporali diverse e per valutare la capacità complessante (CC) si è scelto l'europio (III) come metallo-modello.[2] La determinazione della CC delle acque è stata eseguita mediante titolazione amperometrica, impiegando come titolante l'Eu(III) e utilizzando la tecnica polarografica DPP con elettrodo a goccia di mercurio (polarografo AMEL 433 interfacciato a computer). La CC varia in modo molto netto lungo la colonna d'acqua ed è quasi sempre relativamente elevata se rapportata alla concentrazione totale dei metalli pesanti analizzati (Zn, Cd, Pb e Cu), segno che la capacità del sistema di formare complessi organometallici è ancora discreta. Solo in alcune campagne, ed in particolare nelle acque di fondo, questo parametro ha mostrato valori bassi, indicando una situazione potenzialmente rischiosa. I risultati complessivamente ottenuti consentono una migliore ed aggiornata comprensione dell'impatto degli inquinanti in questa zona costale particolarmente vulnerabile, fornendo utili informazioni non sempre ottenibili dalle attività di monitoraggio di routine. Bibliografia [1] Cozzi, S., Adami, G., Barbieri, P., Cantoni, C., Catalano, G., Cristiani, F., Fioretto, V., Olivo, P., Purini, R., Raicich, F., Reisenhofer, E. 2004. Matching monitoring and modelling in the Gulf of Trieste (Italy). Marine Pollution Bulletin, 48, 587-592. [2] Norden, M., Ephraim, J.H., Allard, B. 1997. Europium complexation by an aquatic fulvic acid - effects of competing ions. Talanta, 44, 781-786.

Structural abnormality of the 11q23 band (11q23+) bearing the MLL gene translocation (MLL+) is a recurrent chromosome change observed in 3% to 7% of acute lymphoblastic leukemias and in 3% to 4% of acute myeloblastic leukemias. The resolution of conventional cytogenetics (CC) in detecting 11q23 rearrangement is limited when the translocative partner has a telomeric location; furthermore, CC can barely discriminate between true 11q23+/MLL+ and rearrangements clustering within the 11q22 to approximately 25 region without MLL involvement (MLL-). We characterized a series of 378 consecutive patients with adult acute leukemia by using CC, fluorescence in situ hybridization (FISH), and multiplex karyotyping (M-FISH) analysis. Our aim was to define the frequency of cryptic MLL+ cases and the frequency of MLL+ within 11q22 to approximately 25+ cases. As expected, FISH was more sensitive than CC in detecting MLL+ cases, but rather unexpectedly, 9 (45%) of 20 patients with 11q22 to approximately 25+ were MLL-. A better characterization of 11q22 to approximately 25+/MLL- leukemias is relevant for the identification of new, recurrent translocations. Moreover, these cases should be readily distinguishable from 11q23+/MLL+ cases. We recommend that karyotypic analysis always be complemented by molecular or FISH methods to unravel MLL rearrangements.

Front line treatment of acute promyelocytic leukemia (APL) with all-trans retinoic acid (ATRA) and chemotherapy (CHT) results in molecular remission in approximately 95% of patients tested after consolidation. The small fraction of patients with persistence of molecular disease (i.e. those in whom polymerase chain reaction (PCR) is positive for PML/RARalpha) after such therapy are thought to have a dismal prognosis but has not yet been investigated in detail.We analyzed the clinico-biological features at presentation of APL patients who showed PCR-detectable residual disease and compared them to those of patients achieving molecular remission after AIDA induction and consolidation. Furthermore, we report the outcome of patients with molecularly persistent disease treated with salvage therapy.Patients attaining molecular remission (n=650) and patients who tested PCR+ve at the end of consolidation (n=23) were not statistically significantly different as regards median age, white cell and platelet counts, morphologic subtype (M3 or M3v), fibrinogen levels or PML/RARalpha transcript type. As to treatment outcome after salvage therapy, 7 patients were treated before morphologic relapse [3 with chemotherapy and autologous stem cell transplantation (SCT) and 4 with allogeneic SCT], and are alive after 64-118 months. Of 16 patients treated at the time of morphologic relapse, only 2 patients are alive, both of whom received an allogeneic SCT.Our findings indicate that APL patients who are molecularly resistant to the AIDA protocol have no distinguishing features at presentation. Their outcome suggests the need for early therapeutic intervention with aggressive treatment prior to the occurrence of hematologic relapse.