Fusion of NUP214 to ABL1 on amplified episomes in T-cell acute lymphoblastic leukemia.

In T-cell acute lymphoblastic leukemia (T-ALL), transcription factors are known to be deregulated by chromosomal translocations, but mutations in protein tyrosine kinases have only rarely been identified. Here we describe the extrachromosomal (episomal) amplification of ABL1 in 5 of 90 (5.6%) individuals with T-ALL, an aberration that is not detectable by conventional cytogenetics. Molecular analyses delineated the amplicon as a 500-kb region from chromosome band 9q34, containing the oncogenes ABL1 and NUP214 (refs. 5,6). We identified a previously undescribed mechanism for activation of tyrosine kinases in cancer: the formation of episomes resulting in a fusion between NUP214 and ABL1. We detected the NUP214-ABL1 transcript in five individuals with the ABL1 amplification, in 5 of 85 (5.8%) additional individuals with T-ALL and in 3 of 22 T-ALL cell lines. The constitutively phosphorylated tyrosine kinase NUP214-ABL1 is sensitive to the tyrosine kinase inhibitor imatinib. The recurrent cryptic NUP214-ABL1 rearrangement is associated with increased HOX expression and deletion of CDKN2A, consistent with a multistep pathogenesis of T-ALL. NUP214-ABL1 expression defines a new subgroup of individuals with T-ALL who could benefit from treatment with imatinib.

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 IgV(H) mutational status. Sixty-five B-CLL patients were investigated using cytogenetics, interphase fluorescence in situ hybridization (FISH), analysis of IgV(H) 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. IgV(H) 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. IgV(H) 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.

Translocation t(1;6)(p35.3;p25.2): a new recurrent aberration in "unmutated" B-CLL.

Although reciprocal chromosomal translocations are not typical for B-cell chronic lymphocytic leukemia (B-CLL), we identified the novel t(1;6)(p35.3;p25.2) in eight patients with this disorder. Interestingly, all cases showed lack of somatically mutated IgV(H). Clinical, morphological, immunologic, and genetic features of these patients are described. Briefly, the age ranged from 33 to 81 years (median: 62.5 years) and the sex ratio was 6M:2F. Most of the patients (6/8) presented with advanced clinical stage. Therapy was required in seven cases. After a median follow-up of 28 months, five patients are alive and three died from disease evolution. Three cases developed transformation into diffuse large B-cell lymphoma. Translocation t(1;6) was found as the primary karyotypic abnormality in three patients. Additional chromosomal aberrations included changes frequently found in unmutated B-CLL, that is, del(11)(q), trisomy 12 and 17p aberrations. Fluorescence in situ hybridization analysis performed in seven cases allowed us to map the t(1;6) breakpoints to the 1p35.3 and 6p25.2 chromosomal bands, respectively. The latter breakpoint was located in the genomic region coding for MUM1/IRF4, one of the key regulators of lymphocyte development and proliferation, suggesting involvement of this gene in the t(1;6). Molecular characterization of the t(1;6)(p35.3;p25.2), exclusively found in unmutated subtype of B-CLL, is in progress.

Megakaryoblastic leukemia with an N-ras mutation and late acquisition of a Philadelphia chromosome.

A patient is described with de novo acute non-lymphocytic leukemia of megakaryoblastic lineage with tri-lineage myelodysplasia. This patient was studied cytogenetically and using molecular genetic techniques throughout her clinical course. She had an N-ras mutation at diagnosis which persisted despite a bone marrow transplant, and acquired a Philadelphia chromosome associated with a P190 BCR-ABL transcript at clinical relapse 3 months post-transplantation.

Variant t(2;11)(p11;q13) associated with the IgK-CCND1 rearrangement is a recurrent translocation in leukemic small-cell B-non-Hodgkin lymphoma.

Classical t(11;14)(q13;q32) involving IGH-CCND1 is typically associated with aggressive CD5-positive mantle cell lymphoma (MCL). Recently, we identified the IGK variant of this translocation, t(2;11)(p11;q13), in three patients with a leukemic small-cell B-non-Hodgkin lymphoma. In all cases, rearrangements of the IGK and CCND1 genes were demonstrated by fluorescence in situ hybridization. Moreover, we mapped the 11q13 breakpoint of this variant translocation in the 3' region of CCND1 which contrasts with the 5' breakpoints in a standard t(11;14)(q13;q32). Expression of cyclin D1 was shown in two cases analyzed either at diagnosis or during disease progression. All three patients were asymptomatic at presentation and no initial therapy was required. One patient died of a progressive disease 58 months from diagnosis, and two patients showed stable disease after 12 months of follow-up. In two analyzed cases, mutated IGVH genes were identified. Our findings indicate that variant t(2;11)(p11;q13) does not typify a classical MCL but possibly a more indolent leukemic lymphoma originating from an antigen experienced (mutated) B cell.

Translocation (11;15)(q23;q14) in three patients with acute non-lymphoblastic leukemia (ANLL): clinical, cytogenetic and molecular studies.

We report on three patients with acute non-lymphoblastic leukemia (ANLL) displaying the same chromosomal translocation t(11;15)(q23;q14). The clinical course of the disease was aggressive, and survival was short. The FAB subtype was M-2 in two cases, and M-1 in the remaining patient. Immunologically two cases showed aberrant expression of a lymphoid antigen (CD19 and TdT, respectively). HTRX1/MLL gene was rearranged in one patient studied at the time of diagnosis. These results plus data scattered in the literature show that the t(11;15)(q23;q14) can be added to the list of recurrent rearrangements in ANLL involving 11q23.

Primary diffuse large B cell lymphoma of the stomach: analysis of somatic mutations in the rearranged immunoglobulin heavy chain variable genes indicates antigen selection.

Gastric low grade MALT lymphomas show a pattern of somatic mutations in their rearranged immunoglobulin genes, indicative of antigen selection. This provides evidence for antigen stimulation in the lymphomagenesis. Gastric diffuse large B cell lymphomas develop secondary to low grade MALT lymphoma or de novo. To study whether antigen-selection is also a feature of primary diffuse large B cell lymphomas, we analysed somatic mutations in the rearranged immunoglobulin heavy chain (IgH) variable genes (VH). The rearranged VH genes of six cases of gastric primary diffuse large B cell lymphoma were amplified from genomic or complementary DNA by a VH gene family-specific polymerase chain reaction method. The PCR products were directly sequenced and were compared to published germline sequences to analyse somatic mutations. Similarly to low grade MALT lymphomas 5/6 primary diffuse large B cell lymphomas show a pattern of somatic mutation in their rearranged VH genes, indicative of antigen selection and suggesting a role for antigens in lymphomagenesis. One case showed bi-allelic VH gene rearrangements, which were non-functional due to extensive deletions. Antigen selection could not be demonstrated or excluded. Antigen selection is a common feature in most analysed primary diffuse large B cell lymphomas, although some heterogeneity in the mechanisms involved in the lymphomagenesis of gastric primary diffuse large B cell lymphomas has not been excluded entirely (case 4).

P190 BCR/ABL transcript in a case of Philadelphia-positive multiple myeloma.

Philadelphia positive multiple myeloma is a very rare event and, so far, no molecular data about the involvement of the BCR and C-ABL genes are available. We report here the case of a 64-year-old woman presenting with a typical multiple myeloma and a complex Philadelphia (Ph) chromosome that we investigated at a molecular level using conventional DNA techniques and the polymerase chain reaction (PCR). No rearrangement was observed within the major breakpoint cluster region (M-BCR) although she was found to have a P190 BCR/ABL hybrid transcript using PCR. As far as we know, this is the first description of a P190-type mRNA in a patient with a chronic lymphoid disorder. Since P190 is almost always associated in man with acute forms of hematological malignancies, this suggests that other factors may play a role in determining the phenotype of the disease.

Translocation t(3;22)(q23;q11) in three patients with diffuse large B cell lymphoma.

In our series of 134 patients with a diagnosis of non-Hodgkin's lymphoma (NHL) and clonal chromosomal abnormalities, three were found to show an identical t(3;22)(q28;q11) translocation. All were old patients with isolated lymphadenomegaly and diffuse large noncleaved cell lymphoma. All expressed a B cell immunophenotype, and all entered a complete remission when treated with aggressive chemotherapy. This translocation could, therefore, delineate a particular subtype of diffuse large cell NHL.

5q- anomaly in lymphoid disorders.

Observations made in two patients and a review of the literature confirm the occurrence of a 5q- chromosome anomaly in lymphoproliferative disorders of both T and B cell type. Additional chromosome changes were invariably present and are of the "lymphoid" type. The chromosome morphology of the 5q- is indistinguishable from that found in myeloid disorders.

Use of dual-color interphase FISH for the detection of inv(16) in acute myeloid leukemia at diagnosis, relapse and during follow-up: a study of 23 patients.

The value of dual-color fluorescence in situ hybridization (FISH) for the detection of inv(16), using two contigs of cosmid probes mapping on both sides of the chromosome 16p breakpoint region, was evaluated in 23 acute myeloid leukemias (AML) in different phases of the disease. At diagnosis interphase FISH detected inv(16) in 19/19 (100%) cases with conventional cytogenetics (CC) evident aberration and excluded the rearrangement in two patients with CC suspected inv(16). Moreover, it also identified an associated del(16p) in two patients. At relapse, it revealed the inv(16) in 8/8 (100%) studied cases. These results were concordant with those of reverse transcriptase-polymerase chain reaction (RT-PCR). From 13 patients who obtained at least one complete remission (CR), 31 follow-up samples were analyzed using interphase FISH. Twenty-nine specimens scored negative for inv(16) and two were positive. RT-PCR detected CBFbeta/MYH11 transcripts in four of the nine CR samples analyzed, being more sensitive than interphase FISH. Eight of the 13 patients relapsed at a median time of 6.5 months (range 1-15) from the last negative FISH analysis. Of the two patients with positive FISH in CR, one relapsed soon after. At diagnosis and relapse, interphase-FISH proved to be an effective technique for detecting inv(16) appearing more sensitive than CC. Prospective studies with more frequent controls and possibly additional FISH probes are needed to assess the value of interphase FISH for minimal residual disease (MRD) and relapse prediction.

Immunoglobulin and T cell receptor gene rearrangement patterns in acute lymphoblastic leukemia are less mature in adults than in children: implications for selection of PCR targets for detection of minimal residual disease.

In order to gain insight into immunoglobulin (Ig) and T cell receptor (TCR) gene rearrangements in adult acute lymphoblastic leukemia (ALL), we studied 48 adult patients: 26 with precursor-B-ALL and 22 with T-ALL. Southern blotting (SB) with multiple DNA probes for the IGH, IGK, TCRB, TCRG, TCRD and TAL1 loci revealed rearrangement patterns largely comparable to pediatric ALL, but several differences were found for precursor-B-ALL patients. Firstly, adult patients showed a lower level of oligoclonality in the IGH gene locus (five out of 26 patients; 19%) despite a comparable incidence of IGH gene rearrangements (24 out of 26 patients; 92%). Secondly, all detected IGK gene deletions (n = 12) concerned rearrangements of the kappa deleting element (Kde) to Vkappa gene segments, which represent two-thirds of the Kde rearrangements in pediatric precursor-B-ALL and only half of the Kde rearrangements in mature B cell leukemias. Thirdly, a striking predominance of immature Ddelta2-Ddelta3 cross-lineage recombinations was observed (seven out of 16 TCRD rearrangements; 44%), whereas more mature Vdelta2-Ddelta3 gene rearrangements occurred less frequently (six out of 16 TCRD rearrangements; 38% vs >70% in pediatric precursor-B-ALL). Together these data suggest that the Ig/TCR genotype of precursor-B-ALL is more immature and more stable in adults than in children. We also evaluated whether heteroduplex analysis of polymerase chain reaction (PCR) products of rearranged Ig and TCR genes can be used for identification of molecular targets for minimal residual disease (MRD) detection. Using five of the major gene targets (IGH, IGK, TCRG, TCRD and TAL1 deletion), we compared the SB data and heteroduplex PCR results. High concordance between the two methods ranging from 96 to 100% was found for IGK, TCRG and TAL1 genes. The concordance was lower for IGH (70%) and TCRD genes (90%), which may be explained by incomplete or 'atypical' rearrangements or by translocations detectable only by SB. Finally, the heteroduplex PCR data indicate, that MRD monitoring is possible in almost 90% of adult precursor-B-ALL and >95% of adult T-ALL patients.

Characteristic pattern of chromosomal gains and losses in marginal zone B cell lymphoma detected by comparative genomic hybridization.

Marginal zone B cell lymphoma (MZBCL) represents a distinct subtype of B cell non-Hodgkin's lymphoma, which has been recently recognized and defined as a disease entity. We investigated 25 cases (18 at primary diagnosis and seven during the course of disease) of MZBCL by comparative genomic hybridization (CGH) and compared these results with cytogenetic, fluorescence in situ hybridization (FISH), and Southern blot data. Twenty of the 25 cases (80%) showed gains (total 49) or losses (total 15) of genetic material. In extranodal, nodal, and splenic MZBCL, material of chromosomes 3 (52% of cases), 18 (32%), X (24%), and 1q (16%) was most frequently gained, whereas losses predominantly involved chromosomes 17 (16%) and 9 (12%). High-level amplifications involving the regions 18q21-23 and 18q21-22, respectively, were detected in two cases. Gains of chromosomes 1q and 8q and losses of chromosome 17 or 17p occurred more frequently in relapsed or progressive lymphomas. For all of the frequently affected chromosomes, CGH allowed narrowing of the relevant subregions including 3q21-23, 3q25-29 and 18q21-23. By Southern blot analysis, the BCL2, BCL6, and CMYC proto-oncogenes were found to be a part of the over-represented regions in two cases, one case, and two cases, respectively, with gains involving 18q, 3q or 8q. In 13 cases, CGH revealed chromosomal imbalances which were not detected by cytogenetic analysis but could be confirmed by FISH or Southern blot analysis in all cases investigated. On the other hand, CGH failed to detect trisomy 3, trisomy 18, and deletion 7q in three cases with a low proportion of tumor cells bearing these abnormalities, as shown by interphase FISH. The characteristic pattern of chromosomal gains and losses detected in this study confirms the distinct nature of MZBCL and may point to chromosomal regions involved in the pathogenesis of these neoplasms.

Hyperthyroidism influences the distribution and apolipoprotein A composition of the high density lipoproteins in man.

Hyperthyroidism has a different influence on the major high density lipoprotein (HDL) components cholesterol, apoprotein (apo) A-I, and apo A-II. To characterize in greater detail the alterations induced by hyperthyroidism within the HDL subclasses, we investigated HDL distribution and composition in 11 hyperthyroid women before and during treatment. The plasma concentrations of total cholesterol, HDL cholesterol, phospholipids, apo A-I, and apo B were decreased when the patients were hyperthyroid compared with the values during treatment. Apo A-II and apo C-III levels were only slightly lower in the hyperthyroid state. Triglyceride and apo E concentrations did not change significantly during therapy. Analysis of lipoprotein subclasses separated by isopycnic ultracentrifugation revealed 1) marked decreases in low density lipoprotein (LDL) cholesterol, phospholipids, and apo B; 2) less pronounced reductions in the very low density lipoprotein (VLDL) lipid and apo B concentrations; and 3) a consistent decrease in the HDL2b (density, 1.063-1.100 g/ml) fraction in the hyperthyroid patients. The reduction in HDL2b mass was associated with lower concentrations of HDL2b cholesterol, phospholipids, and apo A-I. The HDL2b apo A-II levels remained constant during treatment. Hyperthyroidism, therefore, modified the apo A composition of the HDL2b particles and resulted in a decreased molar apo A-I to apo A-II ratio within HDL2b. Further analysis of HDL particles differing in their apo A composition; i.e. HDL particles containing apo A-I only [(A-I)HDL] or containing both apo A-I and A-II [(A-I + A-II)HDL], by immunological procedures suggested that hyperthyroidism influenced the apo A content of HDL2b mainly by changing the proportions of (A-I)HDL and (A-I + A-II)HDL and the amount of apo A-I associated with (A-I)HDL. Treatment reversed the preferential decrease in (A-I)HDL within the HDL2b subclass. The particle sizes within HDL subfractions, measured by polyacrylamide gradient gel electrophoresis, were similar in the untreated and treated patients. Consequently, the decreased mass of apo A-I and lipids within HDL2b in the hyperthyroid patients could be attributed to a reduced number of identically sized particles within this fraction. These data demonstrate that the thyroid hormones are important regulators of HDL metabolism through their influence on the concentration and distribution of apo A-I.

Use of c-KIT/PDGFRA mutational analysis to predict the clinical response to imatinib in patients with advanced gastrointestinal stromal tumours entered on phase I and II studies of the EORTC Soft Tissue and Bone Sarcoma Group.

Previous studies have shown that activating mutations of c-KIT/PDGFRA, potential therapeutic targets for imatinib mesylate, are implicated in the pathophysiology of gastrointestinal stromal tumours (GISTs). In this study, GISTs from 37 patients enrolled in an European Organisation for Research and Treatment of Cancer (EORTC) phase I/II clinical study of imatinib were examined for mutations of c-KIT/PDGFRA in order to explore whether the mutational status of the tumour predicts the clinical response to therapy. Mutations were screened by denaturing high-pressure liquid chromatography (DHPLC) and characterised by bi-directional DNA sequencing. Activating mutations of c-KIT or PDGFRA were found in 29 (78%) and 2 (6%) GISTs, respectively. Most c-KIT mutations involved exon 11 (n=24; 83%), all but one being an in-frame deletion; no isolated point mutations were found. The other c-KIT mutations included exon 9 AY 502-503 duplication (n=4; 14%) and exon 13 Lys-->Glu(642) missense mutation (n=1; 3%). Two tumours with no detectable c-KIT mutations demonstrated PDGFRA Asp-->Glu(842) amino acid substitutions. Patients with GISTs harbouring exon 11 mutations were more likely to achieve a partial response (PR) on imatinib therapy (83%) than all of the others (23%). The overall survival and progression-free survival rates for the entire group at 106 weeks were 78.3% and 46.9%, respectively. Based on a Kaplan-Meier analysis, patients with GISTs harbouring c-KIT mutations had longer median survival times and were less likely to progress than the other patients. These findings indicate that the mutational status of the c-KIT/PDGFRA oncoproteins could be useful to predict the clinical response of patients imatinib therapy.

Histiocyte-rich B-cell lymphoma. A distinct clinicopathologic entity possibly related to lymphocyte predominant Hodgkin's disease, paragranuloma subtype.

This study reports six non-Hodgkin's lymphoma cases that we called histiocyte-rich B-cell lymphoma (BCL) because of the prominent reactive histiocytic infiltrate obscuring the malignant B-cell population. The involved lymph nodes are characterized by a mixed nodular and diffuse infiltrate and occasionally feature prominent sinuses. The infiltrate is composed of reactive lymphocytes and numerous histiocytes obscuring a tumor population composed of variably sized scattered cells with irregular or multilobar vesicular nuclei. Immunostaining of paraffin sections for the B-cell marker recognized by L26 helps in the identification of these neoplastic cells. The clonal nature and further evidence of the B-cell lineage of this condition is shown by immunoglobulin gene rearrangements detected in three cases. The six cases of histiocyte-rich BCL are remarkably similar clinically: all presented with stage IVB disease with splenomegaly and follow an aggressive clinical course. Except for these features, our series show striking similarities to paragranuloma lymphocyte-predominant Hodgkin's disease, including male preponderance (all patients are male), age distribution (mean age, 41 years), propensity to progress to a diffuse, large B-cell lymphoma (two cases), as well as morphology of the neoplastic B-cell population and expression of Hodgkin's cell markers (Leu-M1 positivity after neuraminidase digestion in three cases, Leu-M1 positivity without neuraminidase digestion in one case, and additional epithelial membrane antigen [EMA] positivity in two cases). Both morphologically and clinically, the present series can be differentiated from other types of infiltrate-rich BCL, such as T-cell-rich BCL. Although additional cases will have to be recognized, histiocyte-rich B-cell lymphoma most likely represents a distinct clinicopathological entity. We speculate that it develops from a subset of B cells that also gives rise to the lymphocytic-histiocytic (L/H) cell, the Hodgkin's cell variant of lymphocyte-predominant Hodgkin's disease, paragranuloma subtype.

Discordances of cytoplasmic immunoglobulin G staining with the immunoperoxidase technic in plasma cells from bone marrow, tonsils, and appendix.

Plasma cell cytoplasmic immunoglobulin was stained using the peroxidase-antiperoxidase technic in Bouin-fixed, paraffin-embedded human tissues from different origins. Bone marrow (BM), tonsils, and appendices were examined. IgA-, IgD-, and IgM-secreting plasmocytes were easily studied using highly diluted rabbit antihuman antisera in all tissues, including BM. IgG plasmocytes showed good stainability in tonsils and appendices, but variable results were obtained in BM. Bone marrow IgG plasmocytes from persons without infection required a tenfold higher concentration of rabbit antihuman IgG than plasmocytes derived from patients with infection. Stainability of BM plasmocytes from patients with infection was equal to BM plasmocytes from myeloma patients. Because the same rabbit antihuman IgG concentration could be applied for staining plasmocytes derived from tonsils and appendices, it is most probable that the difference in staining ability is due to a difference in activity of the plasmocytes, i.e., a different IgG concentration in the plasmocytes.

A translocation (8;14) in a cutaneous large B-cell lymphoma.

A 62-year-old man with a cutaneous large-cell (cleaved, noncleaved) lymphoma had multiple facial lesions and two enlarged cervical lymph nodes. Cytogenetic analysis performed on the involved skin and lymph node revealed the following karyotype: 47,XY,inv dup(1)(q11-->q32),+3,t(8;14)(q24;q32). A t(8;14)(q24;q32) translocation is the chromosome anomaly in 75% of Burkitt's lymphomas. Other types of non-Hodgkin's lymphomas, however, may show the same translocation, especially large-cell lymphomas. Duplication of material belonging to the long arm of chromosome 1 is the most frequent additional aberration to Burkitt's translocations in Burkitt's lymphoma/leukemia. Trisomy 3 may be seen in both B- and T-cell malignancies, as well as in angioimmunoblastic lymphadenopathy. Immunogenotyping of malignant cells from a lymph node showed rearrangement of the immunoglobulin heavy and lambda light chain genes. Epstein-Barr virus (EBV) serology was positive for EBV nuclear antigen and EBV capsular antigen. No integration of EBV DNA in tumor tissue, however, could be detected by polymerase chain reaction. These results may be useful in defining the biologic characteristics of cutaneous B-cell non-Hodgkin's lymphomas.

Rare HRAS1 alleles outside the VTR region in lymph nodes from patients with malignant lymphoma.

We found a new type of rare HRAS1 alleles arising de novo in the lymph node of lymphoma patients. Rare alleles of 6.2 kb (a0.1) and 7.3 kb (a2.3) are displayed in the presence of common alleles of 6.6 kb (a1) and 7.6 kb (a3), respectively. These rare alleles lack nucleotide sequences, present in the common alleles in the 5' part between the BamHI and XmaIII site. In addition, the rare alleles may have the classic A-G mutation in the intron at position 2719. In contrast to what is observed for rare constitutional alleles, the VTR region of these rare alleles has the same size as the common alleles. At present the significance of this new type of alleles is not clear.

Molecular breakpoints of t(11;14)(q13;q32) in multiple myeloma.

We found a t(11;14)(q13;q32) translocation in six patients with multiple myeloma (MM) or plasma cell leukemia. In five of them, rearrangements of BCL1 and PRAD1 could be studied. Two patients showed a rearrangement only with the Prad1 probe, located 120 kb telomeric of the major translocation cluster. The three other patients lacked rearrangements with both Bcl1 and Prad1.