Chromothripsis is a frequent event and underlies typical genetic changes in early T-cell precursor lymphoblastic leukemia in adults.

Chromothripsis is a mitotic catastrophe that arises from multiple double strand breaks and incorrect re-joining of one or a few chromosomes. We report on incidence, distribution, and features of chromothriptic events in T-cell acute lymphoblastic leukemias (T-ALL). SNP array was performed in 103 T-ALL (39 ETP/near ETP, 59 non-ETP, and 5 with unknown stage of differentiation), including 38 children and 65 adults. Chromothripsis was detected in 11.6% of all T-ALL and occurred only in adult cases with an immature phenotype (12/39 cases; 30%). It affected 1 to 4 chromosomes, and recurrently involved chromosomes 1, 6, 7, and 17. Abnormalities of genes typically associated with T-ALL were found at breakpoints of chromothripsis. In addition, it gave rise to new/rare alterations, such as, the SFPQ::ZFP36L2 fusion, reported in pediatric T-ALL, deletions of putative suppressors, such as IKZF2 and CSMD1, and amplification of the BCL2 gene. Compared to negative cases, chromothripsis positive T-ALL had a significantly higher level of MYCN expression, and a significant downregulation of RGCC, which is typically induced by TP53 in response to DNA damage. Furthermore we identified mutations and/or deletions of DNA repair/genome stability genes in all cases, and an association with NUP214 rearrangements in 33% of cases.

High grade B-cell lymphoma with MYC, BCL2 and/or BCL6 rearrangements: unraveling the genetic landscape of a rare aggressive subtype of non-Hodgkin lymphoma.

High-grade B-cell lymphoma with MYC and BCL2 and/or BCL6 rearrangements (DH/TH-HGBL) still miss an in-depth genomic characterization. To identify accompanying genetic events, we performed a pilot study on 7 cases by applying DNA microarray and targeted NGS sequencing. Interestingly, the genetic background of DH/TH-HGBL is largely overlapping with that of other high-grade/poor prognosis lymphomas. Namely, copy number abnormalities were trisomy of chromosome 7 and chromosome 8q gain, encompassing MYC. Among gene variants, those affecting transcription factors (MYC, FOXO1), epigenetic modulators (KMT2D, EZH2 and CREEBP), and anti-apoptotic gene (BCL2), were recurrent. MYC and BCL2 were mutated in 3 and 5 cases, respectively. In addition, mutations of FOXO1, previously reported in Diffuse Large B-Cell Lymphomas, were also detected. Clarifying the genomic background of this subset of high-risk lymphomas will pave the way for the clinical use of new biomarkers to: (1) monitor treatment response and; (2) consider alternative targeted therapies.

Endoscopic preaponeurotic access for complex ventral hernia repair with sublay mesh and bilateral anterior component separation: A case report.

INTRODUCTION: Minimally invasive techniques are now routine in complex abdominal wall defects repair. Although laparoscopy allows to reduce post-operative pain, promoting a more rapid recovery and shortening hospital stay, it is associated with risk of bowel injury and adhesions development, when intraperitoneal mesh is placed. We report the case of a patient affected by large recurrent incisional hernia, treated with a new hybrid endoscopic approach. PRESENTATION OF CASE: Patient treated with the novel approach is a 53-year-old male, BMI 27, smoker, with epigastric recurrence of incisional hernia and prosthetic fistula. An endoscopic preaponeurotic subcutaneous access was used. Repair with sublay mesh, bilateral anterior component separation and muscular reinsertions was conducted. Three months after surgery, no signs of recurrence were observed and complete functional recovery had been achieved. DISCUSSION: The new technique adopted benefits from all the advantages of minimally invasive surgery, allowing to avoid risks associated with laparoscopic access. Bilateral anterior component separation with muscular reinsertions is the key for tension-free suture. CONCLUSION: To our knowledge, this is the first time that a complex recurrent incisional hernia is repaired with the hybrid technique aforementioned. The approach used is certainly technically challenging, thus requiring a team skilled in the use of laparoscopy. Good outcomes reported are a further demonstration that minimally invasive surgery can be a valid alternative to traditional open techniques for large abdominal wall defects repair.

A NUP98-positive acute myeloid leukemia with a t(11;12)(p15;q13) without HOXC cluster gene involvement.

We report a case of adult acute myeloid leukemia with a new t(11;12)(p15;q13) underlying a NUP98 rearrangement without HOXC cluster gene involvement. We designed a specific double-color double-fusion FISH assay to discriminate between this t(11;12)(p15;q13) and those producing NUP98-HOXC11 or NUP98-HOXC13. Our fluorescence in situ hybridization (FISH) showed that putative candidate partners mapping 600 kilobases centromeric to HOXC were RARG (retinoic acid receptor gamma), MFSD5 (major facilitator superfamily domain containing 5), and ESPL1 (extra spindle pole bodies homolog 1). It is noteworthy that so far only ESPL1 has been implicated in human cancers. This FISH assay is useful for diagnostic screening of NUP98-positive leukemias.

Molecular cytogenetic findings in a four-way t(1;12;5;12)(p36;p13;q33;q24) underlying the ETV6-PDGFRB fusion gene in chronic myelomonocytic leukemia.

Fluorescence in situ hybridization (FISH) and reverse-transcription polymerase chain reaction (RT-PCR) detected the ETV6-PDGFRB fusion in a patient with chronic myelomonocytic leukemia characterized by bone marrow and peripheral blood eosinophilia and a four-way t(1;12;5;12)(p36;p13;q33;q24) on bone marrow cells. The patient consequently underwent imatinib mesylate therapy and achieved hematologic, FISH, and molecular remission. The FICTION technique (fluorescence immunophenotyping and interphase cytogenetics as a tool for the investigation of neoplasms) demonstrated that eosinophils and CD13(+) and CD14(+) cells belong to the neoplastic clone bearing the ETV6-PDGFRB rearrangement. Molecular cytogenetics is the most reliable approach to detect the involvement of promiscuous genes, such as PDGFRB, and to properly classify genetic entities for which targeted therapies are available. Assessment of cell lineages harboring the genomic lesion may contribute in the understanding of leukemogenic pathways.

A common 93-kb duplicated DNA sequence at 1q21.2 in acute lymphoblastic leukemia and Burkitt lymphoma.

In three patients with acute lymphoblastic leukemia (ALL) and in another with Burkitt lymphoma (BL), conventional cytogenetics and fluorescence in situ hybridization (FISH), applied singly or in combination, showed 1q duplication in two cases of ALL with hyperdiploid karyotypes, 1q duplication resulting from an unbalanced translocation in a third case of ALL, and inv dup(1)(q) in a patient with BL. Centromeric or telomeric breakpoints and extension of the 1q duplicons varied in each case. FISH defined a minimal, common duplicated region of 93kb at band 1q21.2 corresponding to clone RP11-212K13. In this region three putative oncogenes or tumor suppressor genes have been mapped: SF3B4 (splicing factor 3b, subunit 4), OTUD7B (OTU domain containing 7B), and MTMR11 (myotubularin related protein 11). For the first time, a minimal common 1q21.2 duplicated sequence has been identified in lymphoid malignancies in a region where putative oncogenes or suppressor genes have been mapped. This finding elucidates the genomic background of ALL and BL with 1q duplication and provides the basis for molecular studies investigating which genes are involved in leukemogenesis or disease progression in these cases.

Chronic lymphocytic leukaemia Is terminal del(14)(q24) a new marker for prognostic stratification?

In a patient with chronic lymphocytic leukemia (CLL) molecular cytogenetics showed terminal del(14)(q24). Fluorescence in situ hybridisation (FISH) narrowed the deletion to a 35 megabases DNA segment, with the proximal breakpoint between two partially overlapping clones, RP1-116J24 and RP5-1119N5. Besides loss of material at 14q24-qter, comparative genomic hybridisation (CGH) showed loss of 3p21.3-pter, 4p11-p15.1, 8p12-pter, 13q12-q14, and 15q11-q15, and gain of 3q25-qter. Del(13)(q12-14) included the RB-1 gene but not D13S319 and D13S25 loci. The patient was refractory to fludarabine and rituximab. Our findings and data from other reports suggest del(14)(q24) is indicative of aggressive course and is closely associated with del(13)(q14) in CLL.

Submicroscopic deletions in 5q- associated malignancies.

BACKGROUND AND OBJECTIVES: The deletion of the long arm of chromosome 5 is common in myelodysplastic syndromes (MDS) but is not limited to the 5q- syndrome as it is also seen in acute myeloid leukemia (AML), where it is often associated with other karyotypic aberrations. The aim of this study was to investigate whether deletions of known suppressor sequences occur in myeloid malignancies associated with 5q-. DESIGN AND METHODS: Thirty patients with MDS or AML were selected for the presence of a 5q karyotypic deletion, either isolated (19 cases) or associated with other chromosome changes (11 cases). Multiple fluorescent in situ hybridization (FISH) in interphase nuclei was applied in all cases using a panel of eleven probes for known suppressor genes or loci deleted in MDS/AML. Metaphase FISH was also performed to clarify discrepancies between conventional and molecular cytogenetics. RESULTS: No additional deletions were found in nineteen cases with an isolated 5q-. Mono-allelic deletions where found in 9/11 cases, 3 of which were related to monosomies by conventional cytogenetics. Interphase-FISH showed p53, AML1, D13S25, NF1, or Ikaros in six out of nine (66%) patients with 5q- and additional karyotypic changes. Metaphase FISH was helpful in assigning some of these cryptic events to non-proliferating cells. INTERPRETATION AND CONCLUSIONS: Our study emphasizes that isolated 5q- is the marker of a highly stable clone in both MDS and AML. AML with isolated 5q- are molecularly closer to 5q- syndrome than to AML with complex changes. Interphase-FISH data strongly support a mutator phenotype underlying complex karyotypes with a 5q deletion.

Different mechanisms lead to a karyotypically identical t(20;21) in myelodysplastic syndrome and in acute myelocytic leukemia.

A new t(20;21)(q11;q11), associated with a deletion on the long arm of chromosome 20, was found in one patient with an acute myelocytic leukemia (AML) and in one with myelodysplastic syndrome (MDS). In both cases deletion was interstitial, extending from band q11 to band q13, as shown by comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH). FISH analysis with whole arm paints, subtelomeric probes, and locus-specific probes for the long arms of chromosomes 20 and 21 revealed in patient 1 a reciprocal translocation between the deleted 20q and the long arm of chromosome 21, that is, del(20)(q11q13)t(20;21)(q11;q11), and in patient 2, material from 21q was inserted into the deleted 20q, that is, del(20)(q11q13)ins(20;21)(q11;q11q22). This is the first identification of a complex 20;21 rearrangement in MDS/AML. Deletion at 20q and juxtaposition between 20q11 and 21q11 appear to be the critical genomic events.

Interpretation of the complex karyotype and identification of a new 6p amplicon by integrated comparative genomic hybridization and fluorescence in situ hybridization on the U937-I cell line.

Molecular cytogenetics is helpful to identify complex and cryptic genomic changes in malignancy. Human leukemic cell lines are an important tool for advancements of biological research on malignant cells, one critical step being characterization of genomic changes. We used fluorescence in situ hybridization and comparative genomic hybridization to refine karyotypic interpretation of the diffuse histiocytic lymphoma derived U937-1 cell line. From this integrated approach, chromosome material involved in nine karyotypic markers and in unbalanced translocations could be identified. Moreover, a previously undetected amplicon emerged within band 6p21. The U937-I is a new in vitro model to study genome amplification and unknown recombinations in leukemic cells, such as those involving the centromeric region of chromosome 1.