14q32 rearrangements deregulating BCL11B mark a distinct subgroup of T-lymphoid and myeloid immature acute leukemia

Acute leukemias (ALs) of ambiguous lineage are a heterogeneous group of high-risk leukemias characterized by coexpression of myeloid and lymphoid markers. In this study, we identified a distinct subgroup of immature acute leukemias characterized by a broadly variable phenotype, covering acute myeloid leukemia (AML, M0 or M1), T/myeloid mixed-phenotype acute leukemia (T/M MPAL), and early T-cell precursor acute lymphoblastic leukemia (ETP-ALL). Rearrangements at 14q32/BCL11B are the cytogenetic hallmark of this entity. In our screening of 915 hematological malignancies, there were 202 AML and 333 T-cell acute lymphoblastic leukemias (T-ALL: 58, ETP; 178, non-ETP; 8, T/M MPAL; 89, not otherwise specified). We identified 20 cases of immature leukemias (4% of AML and 3.6% of T-ALL), harboring 4 types of 14q32/BCL11B translocations: t(2,14)(q22.3;q32) (n = 7), t(6;14)(q25.3;q32) (n = 9), t(7;14)(q21.2;q32) (n = 2), and t(8;14)(q24.2;q32) (n = 2). The t(2;14) produced a ZEB2-BCL11B fusion transcript, whereas the other 3 rearrangements displaced transcriptionally active enhancer sequences close to BCL11B without producing fusion genes. All translocations resulted in the activation of BCL11B, a regulator of T-cell differentiation associated with transcriptional corepressor complexes in mammalian cells. The expression of BCL11B behaved as a disease biomarker that was present at diagnosis, but not in remission. Deregulation of BCL11B co-occurred with variants at FLT3 and at epigenetic modulators, most frequently the DNMT3A, TET2, and/or WT1 genes. Transcriptome analysis identified a specific expression signature, with significant downregulation of BCL11B targets, and clearly separating BCL11B AL from AML, T-ALL, and ETP-ALL. Remarkably, an ex vivo drug-sensitivity profile identified a panel of compounds with effective antileukemic activity

NPM1 Deletion Is Associated with Gross Chromosomal Rearrangements in Leukemia

BACKGROUND: NPM1 gene at chromosome 5q35 is involved in recurrent translocations in leukemia and lymphoma. It also undergoes mutations in 60% of adult acute myeloid leukemia (AML) cases with normal karyotype. The incidence and significance of NPM1 deletion in human leukemia have not been elucidated. METHODOLOGY AND PRINCIPAL FINDINGS: Bone marrow samples from 145 patients with myelodysplastic syndromes (MDS) and AML were included in this study. Cytogenetically 43 cases had isolated 5q-, 84 cases had 5q- plus other changes and 18 cases had complex karyotype without 5q deletion. FISH and direct sequencing investigated the NPM1 gene. NPM1 deletion was an uncommon event in the "5q- syndrome" but occurred in over 40% of cases with high risk MDS/AML with complex karyotypes and 5q loss. It originated from large 5q chromosome deletions. Simultaneous exon 12 mutations were never found. NPM1 gene status was related to the pattern of complex cytogenetic aberrations. NPM1 haploinsufficiency was significantly associated with monosomies (p<0.001) and gross chromosomal rearrangements, i.e., markers, rings, and double minutes (p<0.001), while NPM1 disomy was associated with structural changes (p=0.013). Interestingly, in complex karyotypes with 5q- TP53 deletion and/or mutations are not specifically associated with NPM1 deletion. CONCLUSIONS AND SIGNIFICANCE: NPM1/5q35 deletion is a consistent event in MDS/AML with a 5q-/-5 in complex karyotypes. NPM1 deletion and NPM1 exon 12 mutations appear to be mutually exclusive and are associated with two distinct cytogenetic subsets of MDS and AML

Caratterizzazione genotipica di una famiglia con beta-talassemia mediante NGS

Introduzione Sempre più frequentemente si osserva una eterogeneità clinica, in termini di carico trasfusionale, insorgenza di complicanze e risposta alla terapia, in pazienti beta-talassemici con genotipo identico. E’ noto che variazioni geniche concomitanti alla mutazione beta, come quelle a carico dei geni alfa globinici e dei geni regolatori BCL11A, HBG2 e HBS1L-MYB, modulano il fenotipo in alcuni casi. L’analisi NGS offre la possibilità di identificare nuovi geni e nuove variazioni coinvolte nella modulazione del fenotipo. Materiali e Metodi Lo studio è stato condotto su 2 fratelli affetti da Beta Talassemia Major, con identico genotipo beta -29 A&gt;G /cod8 (-AA) e geni alfa normali con diversa espressività fenotipica. Abbiamo sviluppato un pannello NGS (SOPHiA GENETICS; tecnologia Illumina Miseq) per lo studio dei geni HBB, HBA1, HBA2, HBD, BMP6, BMP2, ERFE, FTL, HAMP, HFE, HJV, SLC11A2, SLC40A1, TFR2, TMPRSS6, PKLR, KFL1, BCL11A. L’analisi dei dati è stata effettuata con tre criteri di ammissione: 1) SNP &gt;20% e INDEL &gt;15%, 2) coverage &gt;50X, 3) esclusione delle varianti off target (SOPHiA DDM®). Risultati Sono state identificate 44 varianti nel pt.1 e 45 del pt.2. Di queste quattro varianti sono presenti solo nel pt.1 (ERFE:c.*889G&gt;C in eterozigosi, SLC40A1:c.*1130A&gt;G in eterozigosi, TMPRSS6:c.-235T&gt;C e TMPRSS6:c.15C&gt;T in omozigosi) e due solo nel pt.2 (SLC40A1:c.663T&gt;C in eterozigosi e TMPRSS6:c.72G&gt;A in eterozigosi). Inoltre 9 varianti comuni (1 in ERFE, 2 in BCL11A, e 6 in TMPRSS6) presentavano differenze nell’assetto genotipico (omozigote/eterozigote). Conclusioni La caratterizzazione genomica individuale rappresenta sempre più una necessità in funzione della comprensione della correlazione tra genotipo/fenotipo in ogni singolo paziente e nella sua potenziale stratificazione prognostica e terapeutica. Un approccio NGS consente lo studio di variazioni a carico di numerosi geni potenzialmente coinvolti nella modulazione del fenotipo talassemico. I nostri dati, seppur preliminari, mostrano che 6 varianti segregano diversamente nei due fratelli. I geni più ricorrentemente coinvolti sono SLC40A1 (=2) e TMPRSS6 (=3). Quest’ultimo presenta anche il maggior numero di differenze nell’assetto genotipico

T-Cell Acute Lymphoblastic Leukemia: Biomarkers and Their Clinical Usefulness

T-cell acute lymphoblastic leukemias (T-ALL) are immature lymphoid tumors localizing in the bone marrow, mediastinum, central nervous system, and lymphoid organs. They account for 10–15% of pediatric and about 25% of adult acute lymphoblastic leukemia (ALL) cases. It is a widely heterogeneous disease that is caused by the co-occurrence of multiple genetic abnormalities, which are acquired over time, and once accumulated, lead to full-blown leukemia. Recurrently affected genes deregulate pivotal cell processes, such as cycling (CDKN1B, RB1, TP53), signaling transduction (RAS pathway, IL7R/JAK/STAT, PI3K/AKT), epigenetics (PRC2 members, PHF6), and protein translation (RPL10, CNOT3). A remarkable role is played by NOTCH1 and CDKN2A, as they are altered in more than half of the cases. The activation of the NOTCH1 signaling affects thymocyte specification and development, while CDKN2A haploinsufficiency/inactivation, promotes cell cycle progression. Among recurrently involved oncogenes, a major role is exerted by T-cell-specific transcription factors, whose deregulated expression interferes with normal thymocyte development and causes a stage-specific differentiation arrest. Hence, TAL and/or LMO deregulation is typical of T-ALL with a mature phenotype (sCD3 positive) that of TLX1, NKX2-1, or TLX3, of cortical T-ALL (CD1a positive); HOXA and MEF2C are instead over-expressed in subsets of Early T-cell Precursor (ETP; immature phenotype) and early T-ALL. Among immature T-ALL, genomic alterations, that cause BCL11B transcriptional deregulation, identify a specific genetic subgroup. Although comprehensive cytogenetic and molecular studies have shed light on the genetic background of T-ALL, biomarkers are not currently adopted in the diagnostic workup of T-ALL, and only a limited number of studies have assessed their clinical implications. In this review, we will focus on recurrent T-ALL abnormalities that define specific leukemogenic pathways and on oncogenes/oncosuppressors that can serve as diagnostic biomarkers. Moreover, we will discuss how the complex genomic profile of T-ALL can be used to address and test innovative/targeted therapeutic options

t(3;11)(q12;p15)/NUP98LOC348801 fusion transcript in acute myeloid leukemia

Brief Report In a case of acute myeloid leukemia we report molecular cytogenetic findings of a t(3;11)(q12;p15), characterized as a new NUP98 translocation rearranging with LOC348801 at chromosome 3. NUP98 involvement was detected by fluorescence in situ hybridization. 3’-RACE-PCR showed nucleotide 1718 (exon 13) of NUP98 was fused in-frame with nucleotide 1248 (exon 2) of LOC348801. RT-PCR and cloning experiments detected two in-frame spliced NUP98-LOC348801 transcripts and the reciprocal LOC348801-NUP98. A highly specific double-color double-fusion FISH assay reliably detects NUP98-LOC348801. Key words: acute myeloid leukemia, NUP98, translocation partners

Studio di una famiglia beta talassemica mediante NGS

Introduzione Variazioni geniche concomitanti alle mutazioni beta, (es. geni alfa globinici, geni regolatori BCL11A, HBG2, HBS1L-MYB), possono modulare il fenotipo clinico. L’analisi NGS offre la possibilità di identificare nuovi geni e nuove variazioni coinvolte nella modulazione del fenotipo. Materiali e Metodi Lo studio è stato condotto su 2 fratelli affetti da Beta Talassemia Major, diversa espressività fenotipica, identico genotipo beta -29 A&gt;G /cod8 (-AA) e geni alfa normali. Abbiamo sviluppato un pannello NGS (SOPHiA GENETICS) per lo studio dei geni HBB, HBA1, HBA2, HBD, BMP6, BMP2, ERFE, FTL, HAMP, HFE, HJV, SLC11A2, SLC40A1, TFR2, TMPRSS6, PKLR, KFL1, BCL11A. Risultati Sono state identificate 44 varianti nel pt.1 e 45 del pt.2. Quattro varianti sono presenti solo nel pt.1 (1 in ERFE in eterozigosi, 1 in SLC40A1 in eterozigosi, 2 in TMPRSS6 in omozigosi) e due solo nel pt.2 (1 in SLC40A1 in eterozigosi e 1 in TMPRSS6 in eterozigosi). Inoltre 9 varianti comuni (1 in ERFE, 2 in BCL11A e 6 in TMPRSS6) presentavano differenze nell’assetto genotipico (omozigote/eterozigote). Il pt 1 presenta regime trasfusionale più intenso (2 GRC/settimana vs 1 GRC/settimana) senza sovraccarico marziale RMt2* (deferasirox). Il paziente 2 presenta moderato sovraccarico cardiaco ed epatico, ferritinemia elevata (deferasirox cpr rivestite, pregressa idiosincrasia epatica a deferasirox dispersibile e agranulocitosi con deferiprone). Conclusioni La caratterizzazione genomica individuale rappresenta sempre più una necessità per ottenere una correlazione tra genotipo/fenotipo e una potenziale stratificazione prognostica e terapeutica. I nostri dati, seppur preliminari, mostrano che 6 varianti segregano diversamente nei due fratelli