Arjan Buijs

High cereblon expression is associated with better survival in patients with newly diagnosed multiple myeloma treated with thalidomide maintenance

Recently, cereblon (CRBN) expression was found to be essential for the activity of thalidomide and lenalidomide. In the present study, we investigated whether the clinical efficacy of thalidomide in multiple myeloma is associated with CRBN expression in myeloma cells. Patients with newly diagnosed multiple myeloma were included in the HOVON-65/GMMG-HD4 trial, in which postintensification treatment in 1 arm consisted of daily thalidomide (50 mg) for 2 years. Gene-expression profiling, determined at the start of the trial, was available for 96 patients who started thalidomide maintenance. In this patient set, increase of CRBN gene expression was significantly associated with longerprogression-free survival (P = .005). In contrast, no association between CRBN expression and survival was observed in the arm with bortezomib maintenance. We conclude that CRBN expression may be associated with the clinical efficacy of thalidomide. This trial has been registered at the Nederlands Trial Register (www.trialregister.nl) as NTR213; at the European Union Drug Regulating Authorities Clinical Trials (EudraCT) as 2004-000944-26; and at the International Standard Randomized Controlled Trial Number (ISRCTN) as 64455289.

Reconstructing the human hematopoietic niche in immunodeficient mice: opportunities for studying primary multiple myeloma

Interactions within the hematopoietic niche in the BM microenvironment are essential for maintenance of the stem cell pool. In addition, this niche is thought to serve as a sanctuary site for malignant progenitors during chemotherapy. Therapy resistance induced by interactions with the BM microenvironment is a major drawback in the treatment of hematologic malignancies and bone-metastasizing solid tumors. To date, studying these interactions was hampered by the lack of adequate in vivo models that simulate the human situation. In the present study, we describe a unique human-mouse hybrid model that allows engraftment and outgrowth of normal and malignant hematopoietic progenitors by implementing a technology for generating a human bone environment. Using luciferase gene marking of patient-derived multiple myeloma cells and bioluminescent imaging, we were able to follow pMM cells outgrowth and to visualize the effect of treatment. Therapeutic interventions in this model resulted in equivalent drug responses as observed in the corresponding patients. This novel human-mouse hybrid model creates unprecedented opportunities to investigate species-specific microenvironmental influences on normal and malignant hematopoietic development, and to develop and personalize cancer treatment strategies.

Identification of chromosomal abnormalities relevant to prognosis in chronic lymphocytic leukemia using multiplex ligation-dependent probe amplification

B-cell chronic lymphocytic leukemia (CLL) is characterized by a highly variable clinical course. Characteristic genomic abnormalities provide clinically important prognostic information. Because karyotyping and fluorescence in situ hybridization (FISH) are laborious techniques, we investigated the diagnostic efficacy of the more recently developed multiplex ligation-dependent probe amplification (MLPA) technique. MLPA and interphase FISH data of 88 CLL patients were compared for loci encompassing the 13q14 region, chromosome 12, and the ATM (11q22) and TP53 (17p13) genes. We found a perfect correlation, provided that the abnormal clone was present in at least 10-20% of the cells. Because multiple loci and multiple probes per locus were included in the MLPA assay, additional abnormalities not covered by the FISH probes were detected. Furthermore, in 13 cases deletions partly covering the 13q14.3 locus were observed, including three deletions that remained undetected by FISH. All the deletions included the noncoding RNA locus DLEU1 (previously BCMS), which is considered to be the most likely CLL-associated candidate tumor suppressor gene within the 13q14 region. We conclude that MLPA serves as a comprehensive and reliable technique for the simultaneous identification of different clinically relevant and region-specific genomic aberrations in CLL.

Expression pattern of the septin gene family in acute myeloid leukemias with and without MLL-SEPT fusion genes

Septins are proteins associated with crucial steps in cell division and cellular integrity. In humans, 14 septin genes have been identified, of which five (SEPT2, SEPT5, SEPT6, SEPT9, and SEPT11) are known to participate in reciprocal translocations with the MLL gene in myeloid neoplasias. We have recently shown a significant down-regulation of both SEPT2 and MLL in myeloid neoplasias with the MLL-SEPT2 fusion gene. In this study, we examined the expression pattern of the other 13 known septin genes in altogether 67 cases of myeloid neoplasia, including three patients with the MLL-SEPT2 fusion gene, four with MLL-SEPT6 fusion, and three patients with the MLL-SEPT9 fusion gene. When compared with normal controls, a statistically significant down-regulation was observed for the expression of both MLL (6.4-fold; p=0.008) and SEPT6 (1.7-fold; p=0.002) in MLL-SEPT6 leukemia. Significant down-regulation of MLL was also found in MLL-MLLT3 leukemias. In addition, there was a trend for SEPT9 down-regulation in MLL-SEPT9 leukemias (4.6-fold; p=0.077). Using hierarchical clustering analysis to compare acute myeloid leukemia genetic subgroups based on their similarity of septin expression changes, we found that MLL-SEPT2 and MLL-SEPT6 neoplasias cluster together apart from the remaining subgroups and that PML-RARA leukemia presents under-expression of most septin family genes.

Elucidation of a novel pathogenomic mechanism using genome-wide long mate-pair sequencing of a congenital t(16;21) in a series of three RUNX1 -mutated FPD/AML pedigrees

Elucidation of a novel pathogenomic mechanism using genome-wide long mate-pair sequencing of a congenital t(16;21) in a series of three RUNX1 -mutated FPD/AML pedigrees

A ZMYM2-FGFR1 8p11 myeloproliferative neoplasm with a novel nonsense RUNX1 mutation and tumor lysis upon imatinib treatment

The 8p11 myeloproliferative neoplasm (8p11 MPN) is a rare disorder that is molecularly characterized by fusions of diverse partners to the tyrosine kinase receptor gene FGFR1. It can rapidly transform to acute myeloid leukemia. Here we report on a case with a t(8;13)(p11.2;q12.1) ZMYM2-FGFR1 fusion, with massive tumor lysis upon tyrosine kinase inhibition with imatinib. Upon reevaluation, we detected trisomy 21 in addition to the translocation. Sequencing revealed a nonsense c.958C →T RUNX1 mutation both at diagnosis and disease progression, resulting in a p.Arg320X carboxyl-terminal truncated RUNX1 protein. This is the first report on an 8p11 MPN with a trisomy 21 RUNX1 mutation.

Outcome of allogeneic transplantation in newly diagnosed and relapsed/refractory multiple myeloma: long-term follow-up in a single institution

Allogeneic stem cell transplantation (allo‐SCT) has the potential to induce long‐term remission in multiple myeloma (MM), but the role of allo‐SCT in MM is controversial due to the high rate of treatment‐related mortality (TRM). However, although proteasome inhibitors and immunomodulatory drugs have improved the outcome of patients with MM, high‐risk patients still have a very poor prognosis. This indicates the need for new treatment strategies and identification of patients who might benefit from allo‐SCT. We therefore analyzed the outcome of one hundred and forty‐seven patients with MM who received an allo‐SCT at our institution (58 in first line, 89 in relapsed/refractory setting) after a median follow‐up of 88.8 months. For the first‐line setting, median progression‐free survival (PFS) and overall survival (OS) were remarkably good, with a CR rate of 48.3%, median PFS of 30.2 months, and 10‐yr OS of 51%. We found no difference in outcome for patients with high‐risk metaphase cytogenetics or FISH del(13q14), but efficacy in current standard high‐risk patients could not be determined. The outcome in the relapsed/refractory setting was poor, especially in the subgroup of patients relapsing within 18 months after auto‐SCT. Therefore, if applied at all in these patients, improvement of allo‐SCT is needed, focusing on reduction of TRM and more effective immunotherapy.

A girl with an atypical form of ataxia telangiectasia and an additional de novo 3.14 Mb microduplication in region 19q12

A 9-year-old girl born to healthy parents showed manifestations suggestive of ataxia telangiectasia (AT), such as short stature, sudden short bouts of horizontal and rotary nystagmus, a weak and dysarthric voice, rolling gait, unstable posture, and atactic movements. She did not show several cardinal features typical of AT such as frequent, severe infections of the respiratory tract. In contrast, she showed symptoms not generally related to AT, including microcephaly, profound motor and mental retardation, small hands and feet, severely and progressively reduced muscle tone with slackly protruding abdomen and undue drooling, excess fat on her upper arms, and severe oligoarthritis. A cranial MRI showed no cerebellar hypoplasia and other abnormalities. In peripheral blood samples she carried a de novo duplication of 3.14 Mb in chromosomal region 19q12 containing six annotated genes, UQCRFS1, VSTM2B, POP4, PLEKHF1, CCNE1, and ZNF536, and a de novo mosaic inversion 14q11q32 (96% of metaphases). In a saliva-derived DNA sample only the duplication in 19q12 was detected, suggesting that the rearrangements in blood lymphocytes were acquired. These findings reinforced the suspicion that she had AT. AT was confirmed by strongly elevated serum AFP levels, cellular radiosensitivity and two inherited mutations in the ATM gene (c.510_511delGT; paternal origin and c.2922-50_2940del69; maternal origin). This case suggest that a defective ATM-dependent DNA damage response may entail additional stochastic genomic rearrangements. Screening for genomic rearrangements appears indicated in patients suspected of defective DNA damage responses.

Both SEPT2 and MLL are down-regulated in MLL-SEPT2 therapy-related myeloid neoplasia

BackgroundA relevant role of septins in leukemogenesis has been uncovered by their involvement as fusion partners in MLL-related leukemia. Recently, we have established the MLL-SEPT2 gene fusion as the molecular abnormality subjacent to the translocation t(2;11)(q37;q23) in therapy-related acute myeloid leukemia. In this work we quantified MLL and SEPT2 gene expression in 58 acute myeloid leukemia patients selected to represent the major AML genetic subgroups, as well as in all three cases of MLL-SEPT2-associated myeloid neoplasms so far described in the literature.MethodsCytogenetics, fluorescence in situ hybridization (FISH) and molecular studies (RT-PCR, qRT-PCR and qMSP) were used to characterize 58 acute myeloid leukemia patients (AML) at diagnosis selected to represent the major AML genetic subgroups: CBFB-MYH11 (n = 13), PML-RARA (n = 12); RUNX1-RUNX1T1 (n = 12), normal karyotype (n = 11), and MLL gene fusions other than MLL-SEPT2 (n = 10). We also studied all three MLL-SEPT2 myeloid neoplasia cases reported in the literature, namely two AML patients and a t-MDS patient.ResultsWhen compared with normal controls, we found a 12.8-fold reduction of wild-type SEPT2 and MLL-SEPT2 combined expression in cases with the MLL-SEPT2 gene fusion (p = 0.007), which is accompanied by a 12.4-fold down-regulation of wild-type MLL and MLL-SEPT2 combined expression (p = 0.028). The down-regulation of SEPT2 in MLL-SEPT2 myeloid neoplasias was statistically significant when compared with all other leukemia genetic subgroups (including those with other MLL gene fusions). In addition, MLL expression was also down-regulated in the group of MLL fusions other than MLL-SEPT2, when compared with the normal control group (p = 0.023)ConclusionWe found a significant down-regulation of both SEPT2 and MLL in MLL-SEPT2 myeloid neoplasias. In addition, we also found that MLL is under-expressed in AML patients with MLL fusions other than MLL-SEPT2.

Array CGH in Human Leukemia: From Somatics to Genetics

During the past decade, array CGH has been applied to study copy number alterations in the genome in human leukemia in relation to prediction of prognosis or responsiveness to therapy. In the first segment of this review, we will focus on the identification of acquired mutations by array CGH, followed by studies on the pathogenesis of leukemia associated with germline genetic variants, phenotypic presentation and response to treatment. In the last section, we will discuss constitutional genomic aberrations causally related to myeloid leukemogenesis.