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Background: ALL is a disease characterised by lack of differentiation and uncontrolled proliferation of lymphocyte precursor cells, driven by activation of oncogenes and inactivation of tumour suppressor genes. About 30% of B-cell ALL carries the Philadelphia chromosome (Ph), originated by the translocation t(9;22)(q34;q11), and thus express the fusion gene BCR-ABL. The constitutively active tyrosine kinase encoded by BCR-ABL blocks the differentiation of B precursor cells, prevents apoptosis and also causes genetic instability. This ultimately leads to the acquisition of new mutations, resistance to BCR-ABL inhibitors and disease progression. In particular BCR-ABL stimulates DNA repair by non-homologous end-joining and homologous recombination repair mechanisms after various genotoxic stimuli, but a lot of deletions and abnormalities are also generated. Methods and patients: In order to identify whether some genetic lesions could contribute to genetic instability in Ph+ ALL, we analyzed the genome of 18 Ph+ ALL patients using high-resolution single nucleotide polymorphism (SNP) array. 250 ng of genomic DNA were processed on 250K NspI array according to protocols provided by the manufacturer (Affymetrix Inc., Santa Clara, CA, USA). Copy number state was calculated with respect to a set of 48 Hapmap normal individuals using Partek® Genomics Suite. Fluorescence in situ hybridization (FISH) and real-time quantitative polymerase chain reaction (PCR) were performed to validate our results. Results: We found a significant deletion on chromosome 22, cytoband q11.22, close to immunoglobulin lambda light chain genes, in the region 20913571-20931814, corresponding to the VpreB1 (CD179a, Pre-B-Lymphocyte gene 1) gene in about 30% of newly diagnosed Ph+ ALL patients. It is highly conserved among mammals and selectively expressed during the pre-B stage of B cells ontogenesis and together with IGLL1 encodes for the surrogate light chain of the pre-B cell receptor. In mice its monoallelic deletion is associated with an increase of lymphocytes in pre-B stage in bone marrow, while its complete absence causes the impossibility to generate mature B cells. Since Ph+ ALL cells are immunophenotypically blocked at the pre-B stage, we are investigating whether VpreB1 deletion could interfere with normal B cell development and thus could have a role in leukeamogenesis and disease progression. Conclusion: This study demonstrated that the use of high-resolution SNP array is a powerful technology to detect genetic anomalies not revealed by classical cytogenetics which may provide new insights into leukemogenesis.

Background: The Bcr-Abl kinase expressed in ALL drives malignant transformation of human pre-B cells and promote aberrant alternative splicing of genes involved in B-cell signaling and differentiation. The fact that Ikaros (Ik) transcription factor functions as a critical regulator of normal lymphocyte development and the observation of rapid development of leukaemia in mice expressing non-DNA binding isoforms, prompted our study to investigate whether normal Ik expression and function might be altered in Ph+ ALL. Patients and methods: Bone marrow and peripheral blood samples from 47 adult patients (median age 55 yrs, 18-76) with Ph+ ALL were collected after informed consent. Reverse transcription-polymerase chain reaction (RT-PCR) using specific primers for exon 1 and exon 7 of Ik, cloning, nucleotide sequencing and western blot analyses were performed to identify the specific Ik isoforms. Results: In 43/47 (90%) patients the Ik6 isoform, lacking all N-terminal zinc-fingers responsible for DNA-binding, was detected and in 23 of them (54%) it was the predominant isoform. Ik6 expression strongly correlated with the BCR-ABL transcript levels suggesting that this alteration could depend on the Bcr-Abl activity. Patients expressed dominant-negative Ik6 at diagnosis and at the time of relapse, but never during remission. In the patients who seemed to express by RT-PCR only full-length Ik isoforms cloning and subsequent sequencing revealed clones harboring different transcripts. We detected an in frame deletion of 10 aa (ΔKSSMPQKFLG) at the end of ex6 due to the selection of an alternative donor splice site. The consequences were impaired activation of transcription and impaired ability to form dimers. We also observed isoforms with a 60-bp insertion (TYGADDFRDFHAIIPKSFSR) immediately upstream of exon 4 at the exon 2/exon 4 junction either alone or together with the in-frame deletion. Also in this in case the aberration is due to the selection of alternative splice site which determines the skipping of exon 3 and the insertion of a sequence inside the intron 3-4 which starts with an alternative acceptor splice site. Aberrant Ik isoforms had a cytoplasmatic localization as revealed by immunofluorescence and confocal laser scanning microscopy analyses. Conclusions: Our data demonstrated that aberrant splicing of Ikaros represents a frequent feature in Ph+ ALL patients and that dominant negative Ik6 splice variant correlates with BCR-ABL mRNA levels, disease progression and relapse.

In this study we constructed a scale that measures centrality, social, communal, and interdependent identifi cation, and investigated the distinction between these four different types of identifi cation with social groups. The general aim was to examine the psychometric properties of the newly designed Centrality, Social, Communal, and Interdependent Identifi cation Scale (CSCIIS) and to investigate whether differences in self-construal, relationship orientation, gender, and culture might predict each type of identifi cation. The results provided initial support for the validity and the reliability of the CSCIIS, revealed cross-cultural differences in ingroup identifi cation, and supported predictions regarding the correlations between particular types of relationships orientation and particular types of identifi cation with social groups., Настоящото емпирично изследване се фокусира върху конструирането и валиди- зацията на скала, която измерва централност, социална, общностна, и взаимозависима идентификация и има за цел да направи разграничение между тези четири различни типа идентификация със социални групи. Основната идея е да се проверят психометричните свойства на новосъздадената Скала за централност, социална, общностна и взаимозависи- ма идентификация (СЦСОВИ) и да се установи дали различия в себеконструирането, ори- ентацията към междуличностни взаимоотношения, пола и културата могат да предскажат всеки един тип идентификация. Резултатите предоставят първоначална подкрепа за ва- лидността и надеждността на СЦСОВИ, разкриват междукултурни различия във вътреш- ногруповата идентификация и подкрепят очакванията ни за корелация между определени видове ориентация към междуличностни взаимоотношения и точно определени типове идентификация със социални групи.

Background: Detection of residual leukemic cells by measuring BCR-ABL1 transcript level with assays based on quantitative polymerase chain reaction (qPCR) is necessary in the monitoring of minimal residual disease in patients with BCR-ABL1 positive ALL. Aim: To investigate the efficacy of a high sensitive method based on nanofluidic platform (Fluidigm Corporation, South San Francisco, CA) to detect and quantify residual and rare BCR-ABL1 copies, we compared results obtained by conventional qPCR with those obtained by the nanofluidic approach. Patients and methods: We analyzed 60 BCR-ABL1 positive ALL samples expressing p190 (42) or p210 (18) isoform, first by TaqMan absolute qPCR (Applied Biosystems 7900HT Fast Real-Time PCR System). BCR-ABL1 target gene and ABL1 control gene copies were derived by the interpolation of cycle threshold values to the appropriate standard curve obtained using serial plasmid dilutions containing known BCR-ABL1 or ABL1 gene copies. Results: Samples resulted in complete (22) or major (38) molecular response (BCR-ABL1/ABL1 ratio ≪ 0.001 or < 0.1, respectively; 3,8 BCR-ABL1 mean copy number copies). Subsequently we reanalyzed the samples using the 12.765 Digital array (Fluidigm) and the integrated BioMark Digital PCR Analysis software (Fluidigm). The nanofluidic biochip consists in twelve panels, each containing 765 individual reaction chambers where samples are portioned prior to qPCR. As fluorescent signal is produced only in chambers containing copies of the target sequence, digital array provides an absolute quantification by counting the number of positive reactions. Sensitivity and reproducibility of the assay were assessed using six serial dilutions of plasmids (Ipsogen) expressing known BCR-ABL1 p190 transcript copy number (10000; 1000; 100; 50; 10; 1 copies). A 2 µl volume of input cDNA was loaded and two panel for each dilution were used. Results showed a detection rate until a copy of target sequence and a pairing significantly effective between replicates. We then analyzed duplicates of BCR-ABL1 ALL samples: digital array resulted positive in 21 of major molecular response samples (2,13 BCR-ABL1 mean copy number copies) and in any complete molecular response samples. In conclusion, the Fluidigm nanofluidic platform provides a high sensitive assay, able to detect until a single copy of BCR-ABL1 transcript, and it could provide an accurate monitoring method for BCR-ABL1-positive ALL CR patients. Supported by European LeukemiaNet, AIL, AIRC, FIRB 2006, PRIN 2008, Ateneo RFO grants, Project of integreted program (PIO), Programma di Ricerca Regione – Università 2007 – 2009.

Background. Nelarabine (N) is approved for the treatment of T-ALL and T-LBL that have not responded to or has relapsed after treatment with at least 2 chemotherapy regimens. Aim. To evaluate safety profile and efficacy of N as savage therapy in 16 adult relapsed or refractory T-ALL or T-LBL. Methods. After obtaining an informed consent, 16 patients (median age 33 years, range 19-45, M/F = 13/3) affected by T-ALL (n=10) and T-LBL (n=6) received savage therapy with N (median cycle=1, range 1-3), administered at standard adult dosage (1500 mg/sqm on days 1, 3 and 5, every 21). Four patients were primary resistant to induction treatment, 7 patients were relapsed after two previous chemotherapy regimens (including allogeneic BMT in 4 cases and autologous SCT in 1 case); the remaining 6 patients had a molecular relapsed disease (MRD positive). Molecular characterization was performed, including NOTCH and WT-1 genes mutational status. GEP analysis, according to Ferrando A. stratification (Cancer Cell 2002), is still ongoing. Results. Currently, 12 out of 16 patients are evaluable, due to a too short follow up in the other 4 cases. Seven out of 12 patients obtained a complete remission (CR) (5 T-ALL 2 T-LBL);a partial remission (PR) was documented in 2 cases, with an overall response rate (ORR) of 75%. Median duration of CR was 10 weeks (range 2.8-54+). Among these, 2 out of 4 patients in molecular relapse reached a molecular CR and underwent an allogenic BMT (currently in CR after a median follow up of 12 months). Extra- hematological toxicity, not clearly related to the drug, occurred in 3 cases, determining, a complete and irreversible paraplegia, a condition of mental confusion, and a peripheral neuropathy, respectively. Conclusions. N showed a strong efficacy also in cases with low levels of residual disease, in addition to a good safety profile. Neurological toxicity needs to be strictly monitored. Acknowledgments. European LeukemiaNet, AIL, AIRC, Fondazione Del Monte di Bologna e Ravenna, FIRB 2006, PRIN 2008, Ateneo RFO grants, Project of integrated program (PIO), Programma di Ricerca Regione – Università 2007 – 2009.

Background: Recently, a genome-wide analysis using single nucleotide polymorphism (SNP) microarrays, identified genetic alterations targeting PAX5 in over 30% of pediatric acute lymphoblastic leukemia cases (Mullighan et al. Nature 2008). However, the occurrence of PAX5 alterations has not been investigated in adult BCR-ABL1-positive ALL. Aim: To characterize mutations and rearrangements on 9p involving PAX5 in adult BCR-ABL1-positive ALL. Patients and Methods: Eighty-nine patients with de novo adult BCR-ABL1-positive ALL were enrolled into institutional (n = 15) or Gruppo Italiano Malattie Ematologiche Maligne dell'Adulto Acute Leukemia Working Party (n = 74) clinical trials and after obtaining informed consent their genome was analyzed by SNP arrays (Affymetrix 250K NspI and SNP 6.0), FISH, genomic PCR and resequencing. Results: SNP array analysis identified a loss of PAX5 in 29 patients (33%). In all cases the deletion was heterozygous. Four patterns of PAX5 deletion were observed: focal deletion involving only the PAX5 gene in one case (3%); deletions involving only a portion of PAX5 and both telomeric and centromeric genes in 11 cases (38%) with a median size of 310 kb (range: 101 kb-16,395 kb); broader deletions involving the entire PAX5 locus and a variable number of flanking genes in 10 patients (34%) with a median size of 1,999 kb (range: 567 kb-1,208 kb); deletion of all chromosome 9 or 9p in 7 patients (24%). These deletions may result in PAX5 haploinsufficiency or altered gene expression. In one patient the deletion of PAX5 involved only a subset of exons (2-6) resulting in an alternative transcript encoding a prematurely truncated protein lacking key PAX5 functional domains. To investigate the consequences of genomic PAX5 alteration, quantitative PCR (q-PCR) was used: genomic alterations on 9p13.2 lead to a significant down-modulation at the transcript level of Pax5. Normal and deleted PAX5 subgroups were also investigated for point mutations, analyzing each exon by direct sequencing, but we did not found nucleotide substitutions. Conclusion: PAX5 deletions, but not point mutations, are frequent events in adult BCR-ABL1 positive ALL and are the main mechanism of inactivation of PAX5 in BCR-ABL1 positive ALL. Supported by: European LeukemiaNet, AIL, AIRC, FIRB 2006, Strategico di Ateneo, GIMEMA Onlus.

Background: Although the pathogenesis of BCR-ABL1+ ALL is mainly related to the expression of BCR-ABL1, additional genetic lesions are supposed to be involved in its development and progression. Aim: In order to define the full repertoire of leukemia-related mutations, changes in expression profiles and alternative splicing (AS) events, the leukemia transcriptome of a BCR-ABL1+ ALL patient at diagnosis and relapse was sequenced using a Whole Transcriptome Sequencing (RNA-Seq) approach. Methods: Poly(A) RNA from blast cells was used to prepare cDNA libraries for Illumina/Solexa Genome Analyzer. Obtained sequence reads were mapped to the human genome reference sequence (UCSC hg18) to identify single nucleotide variants (SNVs). Reads that showed no match were mapped to a dataset of all possible splice junctions created in silico to identify AS events. The number of reads corresponding to RNA from known exons was also estimated and a normalized measure of gene expression level (RPKM) was computed. Results: RNA-seq analysis generated 13.9 and 15.8 million reads from de novo and relapsed ALL samples, respectively, detecting transcripts from 62% and 64% of human annotated genes. Low RPKM estimates (0.01

Introduction: Experimental evidence obtained in transgenic mice revealed that PML-RARA is necessary but not sufficient for the development of APL, suggesting that additional genetic mutations are also required for its development. Aim: To define whether additional submicroscopic genomic alterations may characterize APL and may be used to better classify the disease by dissection of genomic subsets. Methods: 105 adult patients with acute myeloid leukemia were analyzed. These cases included all French-American-British subtypes, miscellaneous cytogenetic abnormalities and normal karyotype subgroups. Among these, the M3 subtype included 28 patients, representing the 33% of the whole study population. Genomic DNA was isolated from blast cells and applied to Genome-Wide Human SNP 6.0 array (Affymetrix, Santa Clara, CA) following the manufacturer's instructions. Fluorescence in situ hybridization, quantitative PCR and nucleotide sequencing were used to confirm genomic alterations. Results: A wide spectrum of different copy number alterations (CNAs) were identified in all cases and no significant difference in the average number of alterations was detected among different leukemia cytogenetic subgroups except for the complex subgroup, which had an average of 55 CNA/patient. In APL cases an average of 8 CNAs per case (range, 1-24) was found. The macroscopic alterations were rare, confirmed conventional cytogenetics and involved trisomy of chromosome 8 in 3 cases, loss of chromosome 6, loss of chromosome 20 and deletions on chromosome 9 and 7. Microscopic CNAs (< 1.5 Mbps) involved every chromosome at least once and predominantly chromosomes 1, 2, 9, 15 and 17. Genetic gains were more common than losses and their median size was 300 kb (range 0.2- 1.4 Mb). The majority of lesions were not recurrent, being identified in only a single patient. Focal genetic alterations were detected at the breakpoints of t(15;17)(q22;q21) in PML and RARA genes. Hemizygous deletions were identified at at 2q33.3-q34 involving ERBB4 (v-erb-a erythroblastic leukemia viral oncogene homolog 4 avian), at 3p11.2 (EPHA3) and at 12p13 (ETV6). Deletions also affected genes involved in cell regulations as CDKN2A (9p21) and RB1 (13q14.2). Most frequent gains affected the TP73 gene at 1p36.3, the oncogenes MYC and PVT1 at 8q24 (4.33 Mb), PRAME at the 22q11.22. Copy neutral loss of heterozygosity events affected 1p34.2-1p32.3, 10p11.2 (MLLT10), 11p11.2 (WT1, CDKN1C, HRAS). Patients with more than 10 CNAs were associated with a worse prognosis. Conclusions: These data demonstrate that different cooperating events may be involved in the generation of APL. These novel findings may be used to stratify patients according to genomic changes and to facilitate the screening for novel therapeutic targets. Supported by: AIL, AIRC, FIRB 2006, Fondazione del Monte di Bologna e Ravenna, Strategico di Ateneo.

Background. Approximately 30% of adult ALL patients are characterized by the presence of the Philadelphia (Ph) chromosome, which derives from a reciprocal translocation t(9;22)(q34;q11) and results in a chimeric BCR-ABL oncogene. The prognosis of this subset of patients treated with standard therapies, including multi-agent chemotherapy, Imatinib, and allogeneic stem cell transplantation, is still dismal, due to a high risk of relapse. Dasatinib and Nilotinb are second generation TKIs developed to overcome the problem of resistance to Imatinib in relapsed Ph+ leukemias. Design and Methods. We retrospectively evaluated the single center experience on therapy efficacy of Dasatinib, Nilotinib, and experimental third generation TKIs, administered as second or subsequent line of therapy on 25 relapsed Ph+ adult ALL patients. All patients were previously treated with Imatinib. The median age at time of diagnosis was 50 years (range 18-74), 17 patients were male and 8 female. Ten patients presented a BCR-ABL P190 fusion protein and corresponding fusion transcript, the remaining a BCR-ABL P210. Nineteen patients received Dasatinib, 2 patients Nilotinib and the remaining 4 patients were treated with third generation TKIs. Fourteen patients (56%) were in first relapse, and 7 (28%), 3 (12%) and 1 (4%) were in second, third and fourth relapse, respectively. A mutational analysis was performed in all the patients before TKIs (9 wild type, 16 mutated, including T315I) and at the time of subsequent relapse; gene expression profiling, SNPArray (6.0 Affymetrix chip), and Ikaros deletions were also analyzed. Results. 13 out of 25 patients (52%) obtained a haematological response (HR) (11 patients treated with Dasatinib, 1 patient with Nilotinib and 1 patient with a third generation experimental TKI). 10 patients obtained also a cytogenetic response (CyR) and 6 patients a molecular response (MolR). With a median follow up of 10.8 months (range 2-29 months), median duration of HR, CyR and MolR were 117 days (range 14-385 days); progression free survival were 162 days with Dasatinib and 91 days with Nilotinib. Overall survival was 25.8 months. Interestingly, in 6 out of 9 wild-type patients, treated with Dasatinib, the mutational analysis showed the emergence of T315I or F317I mutation at the time of relapse. Conclusion. Second and third generation TKIs represent a valid approach in relapsed Ph+ adult ALL patients; the subsequent relapse is often associated to the emergence of mutation, conferring resistance to TKIs.

Mutations in the Bcr-Abl kinase domain (KD) are often detected at the time of resistance to tyrosine kinase inhibitor (TKI) therapy in chronic myeloid leukemia (CML) and Philadelphia-positive (Ph+) acute lymphoblastic leukemia (ALL) patients (pts). It is still unclear whether and in how many pts low level mutations may already be detectable at the time of diagnosis. We therefore analyzed cDNA samples from 22 newly diagnosed pts with Ph+ ALL (n=13) or chronic phase (CP)-CML (n=9) who subsequently received TKI therapy (imatinib, dasatinib or nilotinib). Screening for low level mutations was performed by cloning the Bcr-Abl KD (a.a. 206-524) in a bacterial vector and sequencing 200 independent clones for each pt. All pts had evidence of aberrant KD sequences. Three to twelve different mutations were detected in each pt. Each mutation was present in two to five independent clones. A total of 105 mutations (including 35 silent, 5 nonsense, and 65 missense mutations) were observed. The vast majority (98/105, 93%) of them have never been reported in association with TKI resistance and are likely not to confer any advantage under TKI selective pressure. Interestingly, 93/105 (89%) mutations were transitions: G>A (n=28), A>G (n=23), C>T (n=22), T>C (n=20). Such a high prevalence of transitions (normally occurring 1.4 times more frequently than transversions) suggests that a specific mechanism generating mutations is active in Ph+ cells. One of the nine CP-CML pt received hydroxyurea for 6 months before starting imatinib therapy. In this pt, high-sensitivity mutation screening was performed again immediately before imatinib start and showed further accumulation of mutations. Eight Ph+ ALL pts and three CML pts subsequently relapsed with evidence of mutations, but only one with a mutation (T315I) that was already detectable at diagnosis. The remaining eleven pts are in persistent remission after a follow up ranging from 12 to 36 months, although four of them were harbouring known imatinib-(H396P, D276G, E355G) or dasatinib-(F317L) resistant mutations at low levels. Our observations suggest that: a) Bcr-Abl KD mutations can probably be found at diagnosis in all CP-CML and Ph+ ALL pts; b) mutations seem to arise randomly and most of them are silent or not conferring any growth advantage under the selective pressure of TKIs; c) generation of mutations seems to be linked to Bcr-Abl-driven genetic instability; d) TKI-resistant mutations present at low levels at diagnosis do not always outgrow and lead to relapse, probably because some of them arise in cell clones with limited self-renewal capacity. This warns against high-sensitivity mutation screening of all CML and Ph+ ALL pts before the start of TKI therapy.

Background: BCR-ABL1-positive ALL is the most frequent and prognostically unfavorable subtype of adult ALL, mainly because of genetic instability. Activation-induced cytidine deaminase (AID) introduces single-strand breaks into target DNA and produces immunediversity by inducing somatic hypermutations and class-switch recombinations in human immunoglobulin genes (Ig). Aim: Since at much lower frequency AID can also target non-Ig genes and may even act as a genome-wide mutator, we investigated whether AID was expressed in BCR-ABL1-positive ALL and in chronic myeloid leukemia (CML) at the time of progression to blast crisis. Patients and Methods: We analyzed 61 adult de novo BCR-ABL1-positive ALL patients (pts) and 60 CML (chronic phase and myeloid/lymphoid blast crisis) pts. AID cDNA, obtained from bone marrow or peripheral blood, was amplified with two pairs of oligonucleotides, the forward primer of each couple conjugated with a fluorescent dye (fluorescein) at its 5#8217; end. PCR products were then loaded on the ABI Prism 3730 DNA Analyzer for automated capillary gel electrophoresis and the results were plotted with the AbiPrism GeneMapper v3.5 software (Applied Biosystems). Results: On the 61 de novo adult BCR-ABL1-positive ALL pts, AID mRNA and protein were detected in 36 (59%); their expression correlated with BCR-ABL1 transcript levels and disappeared after treatment with tyrosine kinase inhibitors at the time of remission. AID expression was also found in lymphoid blast crisis CML (50%), but not in myeloid lineage or in chronic phase CML. Different isoforms of AID were identified: 13/61 (21%) pts expressed the full-length isoform; 19/61 (31%) co-expressed the wild-type and different AID splice variants with deaminase activity (AID#916;E4a, with a 30 bp deletion of exon 4; AID#916;E4, with the exon 4 deletion; AIDins3, with the retention of intron 3-4); 4/61 (7%) expressed the AID#916;E3-E4 isoform without deaminase activity (deletion of exons 2 and 3). To investigate whether AID introduces DNA-single strand breaks, we performed a genome wide analysis by 250K NspI single nucleotide polymorphism (SNP) array (Affymetrix Inc., USA). Patients who expressed wild-type AID had a higher number of alterations compared to AID-negative (median copy number alteration of 14 versus 4. respectively, p

By genome-wide analyses of DNA copy number abnormalities in adult BCR-ABL1-positive ALL patients (pts) using single nucleotide polymorphism (SNP) microarrays (Affymetrix 500K and SNP 6.0), we identified a high frequency of genetic alterations of regulators of B lymphoid development (PAX5, EBF1, IKZF1) and cell cycle (CDKN2A, CDKN2B, BTG1). The most frequent somatic copy number alteration was deletion on 7p12 of IKZF1 (68/106 patients, 64%), which encodes the transcription factor Ikaros required for the earliest stages of lymphoid lineage commitment. Here, we characterized and mapped all breakpoints in IKZF1 gene to recognize that two major deletions occur in BCR-ABL1-positive ALL: type A characterized by loss of exons 4-7 (44%) and due to breakpoints occurring in introns 3 and 7; type B characterized by removal of exons 2-7 (19%) and due to a variable pattern of breakpoints in introns 1 and 7. In two pts we had both \#916;2-7 and \#916;4-7 deletions and in one we identified a deletion of all IKZF1 gene and part of GRB10. A variable number of nucleotides (patient-specific) were inserted at the conjunction and maintained with fidelity at the time of relapse. The extent of both deletions correlated with the expression of dominant-negative isoforms with cytoplasmatic localization. The IKZF1 alteration was also identified in B-negative ALL pts (41%) and at the progression of CML to lymphoid blast crisis (66%), but never in myeloid blast crisis CML and chronic phase CML. Known DNA sequences were mapped along the breakpoint cluster regions including heptamer recombination signal sequences recognized by the RAG enzymes during V(D)J recombination and activation-induced cytidine deaminase (AID) consensus motifs (DGYW/WRCH). Gene expression profiling analysis of pts with IKZF1 deletion vs wild-type pts identified a unique signature characterized by a down-regulation of genes involved in pre-B-cell differentiation (e.g. VPREB1, VPREB3, IGLL3, BLK), demonstrating that genomic IKZF1 alterations have a strong impact on trascriptoma and contribute to an impaired B-cell differentiation. Univariate analysis showed that the IKZF1 deletion is a negative prognostic marker influencing the cumulative incidence of relapse (10.1 months for pts with deletion vs 56.1 months for wild-type pts, p=0.0103) and disease-free survival (DFS) (10.1 months vs 32.1 months, respectively, p=0.0229). The negative prognostic impact of the IKZF1 deletion on DFS was also confirmed by multivariate analysis (p=0.0445). In conclusion, deletion of IKZF1 is an important event in the development of BCR-ABL1 B-progenitor ALL which significantly influences clinical outcome.

The primary goal of the present research was to explore the relationship between adult attachment styles and four different types of identification with social groups. The results confirmed predictions and revealed that particular prototypic attachment styles are associated with an increase in only certain types of ingroup identification. People with secure attachment style had higher social identification than people with a dismissive-avoidant attachment style. Participants with secure attachment style showed higher communal identification than participants who had either a dismissive-avoidant or a fearful-avoidant attachment style. These findings supported the idea that relationship attachment style has an important effect on the way people identify with their social groups and can serve as a predictor of preferred type of ingroup identity.

The present research tested the idea that the ecological impact of intergroup contact on outgroup attitudes can be fully understood only when relative frequency and relative influence of positive and negative contact are considered simultaneously. Participants from five European countries (Austria, the Czech Republic, Germany, Poland and Slovakia; N = 1,276) freely described their contact experiences with people of neighboring nationalities and then reported on their outgroup attitudes. Contact descriptions were coded for positive versus negative valence and for person- versus situation-framing. Consistently across the five participants groups, positive intergroup contact was reported to occur three times more frequently than negative intergroup contact; however positive contact was found to be only weakly related to outgroup attitudes. On the contrary, the less frequent negative (vs. positive) contact was comparatively more influential in shaping outgroup attitudes, especially when negativity was reported around the contact person, rather than the contact situation. This research’s findings reconcile contrasting lines of past research on intergroup contact and suggest that the greater prevalence of positive contact may compensate for the greater prominence of negative contact, thus leading to modest net improvements in outgroup attitudes after intergroup contact.

Background: The Ph chromosome is the most frequent karyotypic aberration in adults with ALL and it represents the single most significant adverse prognostic marker. Despite the encouraging results achieved with imatinib, resistance develops rapidly and is quickly followed by disease progression. Recently, the introduction of array-based analysis of single nucleotide polymorphism (SNP) has allowed the rapid determination of genome-wide allelic information at high density for a DNA sample. Patients and Methods: In order to identify, at submicroscopic level, genetic lesions which can escape standard cytogenetic observations and may contribute to leukemogenesis and resistance, we profiled, until now, the genomes of 18 patients out of 55 Ph+ ALL patients treated in our institute with imatinib or dasatinib. 250 ng of genomic DNA were processed on 500K SNP array according to protocols provided by the manufacturer (Affymetrix Inc., Santa Clara, CA, USA). Copy number state was calculated with respect to a set of 48 Hapmap normal individuals and a diploid reference set of samples obtained from acute leukaemia cases in remission using Partek® Genomics Suite. Fluorescence in situ hybridization (FISH) and real-time quantitative PCR were performed to validate our results. Results: We identified multiple copy number alterations per case, with deletions outnumbering amplification almost 3:1. Lesions varied from loss or gain of complete chromosome arms (trisomy 4, monosomy 7, loss of 9p, 10q, 14q, 16q and gain of 1q and 17q) to microdeletions and microduplications targeting genomic intervals. The common microdeletions included 1p36.21 (PRAMEF), 3q29 (TFCR), 8p23 (DEFB105A), 14q11.2 (DAD1), 16p13.11 (PDXDC1, NTAN1, RRN3), 16p11.2 (SPN) and 19p13.2 (CARM1, SMARCA4). Some genomic alterations were identified in genes regulating B-lymphocyte differentiation, such as PAX5 (n=3), BLNK (n=1) and VPREB1 (n=6), IKZF1 (n=7) and in genes with an established role in leukemogenesis, such as MDS, BTG1, MLLT3, RUNX1. The most frequently affected genes were those controlling G1/S cell cycle transition (e.g. CDKN2A and CDKN2B). Furthermore, many of the deletions detected included genes encoded for phosphatase proteins (e.g. PTPRD, PPP1R9B, PTPN18) and for zinc-finger proteins. It is noteworthy that some lesions felt in regions lacking annotated genes. Discussion: Using high-resolution genome wide approach we showed that multiple sub-microscopic genomic anomalies, which may be suspected to contribute to leukemogenesis and used as targets for existing or novel drugs, frequently occur in Ph+ ALL