Lucienne Michaux

A comprehensive genetic and histopathologic analysis identifies two subgroups of B-cell malignancies carrying a t(14;19)(q32;q13) or variant BCL3-translocation

The biologic and pathologic features of B-cell malignancies bearing a translocation t(14;19)(q32;q13) leading to a fusion of IGH and BCL3 are still poorly described. Herein we report the results of a comprehensive cytogenetic, fluorescence in situ hybridization (FISH), molecular and histopathological survey of a large series of B-cell malignancies with t(14;19) or variant translocations. A total of 56 B-cell malignancies with a FISH-proven BCL3 involvement were identified with the translocation partners being IGH (n=51), IGL (n=2), IGK (n=2) and a non-IG locus (n=1). Hierarchical clustering of chromosomal changes associated with the t(14;19) indicated the presence of two different groups of IG/BCL3-positive lymphatic neoplasias. The first group included 26 B-cell malignancies of various histologic subtypes containing a relatively high number of chromosomal changes and mostly mutated IgVH genes. This cluster displayed three cytogenetic branches, one with rearrangements in 7q, another with deletions in 17p and a third one with rearrangements in 1q and deletions in 6q and 13q. The second group included 19 cases, mostly diagnosed as B-cell chronic lymphocytic leukemia (B-CLL), and characterized by few additional chromosomal changes (e.g. trisomy 12) and unmutated IgVH genes. In conclusion, our study indicates that BCL3 translocations are not restricted to B-CLL but present in a heterogeneous group of B-cell malignancies.

Chromosomal translocations independently predict treatment failure, treatment-free survival and overall survival in B-cell chronic lymphocytic leukemia patients treated with cladribine

Chromosomal translocations represent an important prognostic indicator in B-cell chronic lymphocytic leukemia (B-CLL). However, their value had been neither determined in homogeneously treated patients nor compared to that of IgVH mutational status. Sixty-five B-CLL patients were investigated using cytogenetics, interphase fluorescence in situ hybridization (FISH), analysis of IgVH and of TP53 mutational status before treatment with 2-chloro-2′-deoxyadenosine (CdA). Translocations (n=45) were detected in 42% of the patients, including both balanced (n=12) and unbalanced (n=33) types. IgVH was mutated in 43% of the patients. Patients with translocations were more heavily pretreated (P=0.05), presented with more complex karyotypes (P<0.001), 17p abnormalities and TP53 mutations, and had a higher failure rate (59 vs 21% in patients without translocations, P=0.004). Patients with unbalanced translocations displayed a shorter median treatment-free survival (TFS, 6.9 vs 35.9 months, log rank 22.72, P<0.001) and overall survival (OS, 13.0 vs 68.0 months, log rank 16.51, P<0.001), as compared to patients without translocation. In multivariate analysis, unbalanced translocations were independently associated with therapeutic failure, short TFS and short OS. IgVH mutational status was independently associated with risk of failure and TFS, but not OS. In B-CLL patients treated with CdA, translocations are strong predictors of outcome.

Multiple myeloma--an update on diagnosis and treatment.

Multiple myeloma is a plasma cell (PC) malignancy characterized by the accumulation of monoclonal PCs in the bone marrow and the production of large amounts of a monoclonal immunoglobulin or paraprotein. In the past years, new approaches in the diagnosis and treatment were introduced aiming to identify high‐risk patients who need proper anti‐myeloma treatment. Intensive therapy including autologous hematopoietic stem cell transplantation and the new agents bortezomib, thalidomide, and lenalidomide have improved patients’ responses. Further optimalization of the different treatment schedules in well‐defined patient groups may prolong their survival. Patient stratification is currently based on patient characteristics, extent of myeloma disease, and associated cytogenetic and laboratory anomalies. More and more gene expression studies are introduced to stratify patients and to individualize therapy.

Heterogeneous patterns of amplification of the NUP214-ABL1 fusion gene in T-cell acute lymphoblastic leukemia

Episomes with the NUP214-ABL1 fusion gene have been observed in 6% of T-ALL. In this multicentric study we collected 27 cases of NUP214-ABL1-positive T-ALL. Median age was 15 years with male predominance. Outcome was poor in 12 patients. An associated abnormality involving TLX1 or TLX3 was found in all investigated cases. Fluorescent in situ hybridization revealed a heterogeneous pattern of NUP214-ABL1 amplification. Multiple episomes carrying the fusion were detected in 24 patients. Episomes were observed in a significant number of nuclei in 18 cases, but in only 1–5% of nuclei in 6. In addition, intrachromosomal amplification (small hsr) was identified either as the only change or in association with episomes in four cases and two T-ALL cell lines (PEER and ALL-SIL). One case showed insertion of apparently non-amplified NUP214-ABL1 sequences at 14q12. The amplified sequences were analyzed using array-based CGH.These findings confirm that the NUP214-ABL1 gene requires amplification for oncogenicity; it is part of a multistep process of leukemogenesis; and it can be a late event present only in subpopulations. Data also provide in vivo evidence for a model of episome formation, amplification and optional reintegration into the genome. Implications for the use of kinase inhibitors are discussed.

Phase I/II study of 2-chloro-2'-deoxyadenosine with cyclophosphamide in patients with pretreated B cell chronic lymphocytic leukemia and indolent non-Hodgkin's lymphoma.

Because of their substantial in vitro synergy, we conducted a dose-escalation study of cyclophosphamide (CP) added to 2-chloro-2′-deoxyadenosine (CdA) in patients with previously treated chronic lymphocytic leukemia and non-Hodgkins lymphoma. CdA was given at a fixed dose (5.6u2009mg/m2/day) as a 2-h intravenous (i.v.) infusion, immediately followed by a 1-h i.v. infusion of CP, for 3 days. The initial daily CP dose was 200u2009mg/m2, and was escalated by 100u2009mg/m2 increments in successive cohorts of three to six patients to determine the maximum-tolerated dose (MTD). Additional patients were included at the MTD to extend toxicity and response analysis. Twenty-six patients received 68 cycles of chemotherapy. The MTD of CP after CdA 5.6u2009mg/m2, was 300u2009mg/m2. Acute neutropenia was the dose-limiting toxicity of this regimen, which was otherwise well tolerated. Delivery of repeated cycles was not feasible in eight patients (31%) because of prolonged thrombocytopenia. Severe infections were seen in three of 68 cycles (4%). The overall response rate was 58% (15 of 26; 95% CI, 36–76%), with 15% complete responses and 42% partial responses. These data show the feasibility of the association of CdA with CP. Given the response rate observed, further studies of this regimen are warranted in untreated patients, in particular with chronic lymphocytic leukemia and with Waldenström macroglobulinemia.

Interstitial del(14)(q) involving IGH: a novel recurrent aberration in B-NHL

Deletions of chromosome 14 recurrently observed in B-cell malignancies are particularly frequent in chronic lymphocytic leukemia (CLL), multiple myeloma (MM), diffuse large B-cell lymphoma (DLBCL) and B-cell acute lymphoblastic leukemia.1 To characterize these aberrations, we initially mapped the deletions in 23 leukemia/lymphoma cases using a tiling path chromosome 14-array CGH (aCGH) platform (resolution of approximately 42xa0kb) comprising 838 bacterial artificial chromosome/P1 artificial chromosome (BAC/PAC) clones from the Chori 32K set (www.ensembl.org). Del(14)(q) have been successfully mapped in all 23 cases (Figure 1a) and subdivided into two main categories: (1) involving or (2) not involving 14q32.33. This latter category grouped eight cases with interstitial deletions ranging in size from 17 to 70xa0Mb and distributed along chromosome 14 (q13 ter) and one case with three dispersed deletions of 1.5, 7.5 and 2.5xa0Mb. The first category of del(14)(q) comprised 14 cases showing the common distal breakpoint mapped in the 105.26–105.41xa0Mb region harboring the IGH genes cluster. The size of the deleted region varied in six cases; their proximal breakpoints were bordered by RP11-164H3/95.19xa0Mb, CTD-2053J06/83.56xa0Mb, RP11-340F04/74.66xa0Mb, RP11-350H11/67.94xa0Mb, RP11-769O09/66xa0Mb and RP11-520H13/65xa0Mb. Particularly intriguing was the finding of exactly the same del(14)(q24.1q32.33) covering the region of approximately 36xa0Mb in eight cases (Figure 1b). The proximal border of this deletion was consistently flanked by RP11-35D12 (68.24xa0Mb)( 2) and RP11-720I19 (68.45xa0Mb)( 1), and the distal breakpoint was bordered by RP11-448N05 (105.26xa0Mb)( 1) and RP11-284A08 (105.43xa0Mb)( 2). Given that these terminal BACs are assigned to IGCH and IGVH, respectively, aCGH results were additionally validated by fluorescence in situ hybridization (FISH) with the LSI-IGH break-apart probe. As expected, all eight analyzed cases (as well as the remaining six cases with nonrecurrent IGH-involving deletions) showed a one-fusion one-green signal pattern due to loss of the 3IGH flanking sequences (red signal). The proximal breakpoint of the del(14)(q24.1q32.33) validated with RP11-35D12 and RP11-720I19 showed lost or diminished signal from the latter clone (Figure 1b) and thereby pointed to a breakpoint in the region covered by the overlapping extremities of these BAC clones. The only gene located in this region was ZFP36L1. For FISH detection of the del(14)(q24.1q32.33) in other B-cell non-Hodgkins lymphoma (B-NHL), a dual-color break-apart assay for the 14q24.1 breakpoint (RP11-35D12-SO/RP11-720I19-SG) and LSI-IGH were applied. The screening of 58 B-NHL cases with cytogenetic and/or FISH evidence of del(14)(q) led to identification of 13 additional cases with del(14)(q24.1q32.33), including one with the deletion masked by t(14;22)(q32;q11)/IGH-IGL (case 7). Additional FISH screening of 60 random CLL cases, seven MM cell lines, including four (RPMI-8226, L-363, OPM-1 and LP-1) with a previously described del(14),2 and 20 various B-NHL cases with structural aberrations of 14q21–q24, failed to identify ZFP36L1 and/or IGH rearrangements, indicating that these deletions are rare molecular events.

Myelodysplastic syndrome with monosomy 5 and/or 7 following therapy with 2-chloro-2'-deoxyadenosine.

A few cases of secondary neoplasms occurring after treatment with 2‐chloro‐2′‐deoxyadenosine (2CdA) have been reported, mostly in patients previously exposed to other anti‐cancer drugs including alkylating agents (AA). Here we report on the occurrence of a myelodysplastic syndrome (MDS) with monosomy 5 and/or 7 in two patients after 2CdA treatment, without or prior to other toxic exposure. In light of a literature review and given the involvement of chromosomes frequently abnormal in secondary leukaemias, we suggest that 2CdA may induce therapy‐related MDS (t‐MDS).

RPL5 on 1p22.1 is recurrently deleted in multiple myeloma and its expression is linked to bortezomib response

Chromosomal region 1p22 is deleted in ⩾20% of multiple myeloma (MM) patients, suggesting the presence of an unidentified tumor suppressor. Using high-resolution genomic profiling, we delimit a 58u2009kb minimal deleted region (MDR) on 1p22.1 encompassing two genes: ectopic viral integration site 5 (EVI5) and ribosomal protein L5 (RPL5). Low mRNA expression of EVI5 and RPL5 was associated with worse survival in diagnostic cases. Patients with 1p22 deletion had lower mRNA expression of EVI5 and RPL5, however, 1p22 deletion status is a bad predictor of RPL5 expression in some cases, suggesting that other mechanisms downregulate RPL5 expression. Interestingly, RPL5 but not EVI5 mRNA levels were significantly lower in relapsed patients responding to bortezomib and; both in newly diagnosed and relapsed patients, bortezomib treatment could overcome their bad prognosis by raising their progression-free survival to equal that of patients with high RPL5 expression. In conclusion, our genetic data restrict the MDR on 1p22 to EVI5 and RPL5 and although the role of these genes in promoting MM progression remains to be determined, we identify RPL5 mRNA expression as a biomarker for initial response to bortezomib in relapsed patients and subsequent survival benefit after long-term treatment in newly diagnosed and relapsed patients.

CLONAL MONOSOMY 7 AND 5q IN A CHILD WITH MYELODYSPLASTIC SYNDROME

The authors report the case of a 5-year-old boy referred for thrombocytopenia and neutropenia. Bone marrow examination showed a myelodysplasia with clonal monosomy7. The acceleration of the disease was marked by the appearance of an additional cytogenetic abnormality, i.e., the deletion of the long arm of chromosome 5 in the clonal cells. RAS genemutationwas not detected. Chemotherapy was started to achieve complete remission before a bone marrow transplatation. This treatment was complicated by a prolonged a plasia and the patient died of systemic mycotic infection.