Chromosome translocations in multiple myeloma

Multiple myeloma (MM), a malignant tumor of somatically mutated, isotype-switched plasma cells (PC), usually arises from a common benign PC tumor called Monoclonal Gammopathy of Undetermined Significance (MGUS). MM progresses within the bone marrow, and then to an extramedullary stage from which MM cell lines are generated. The incidence of IgH translocations increases with the stage of disease: 50% in MGUS, 60-65% in intramedullarly MM, 70-80% in extramedullary MM, and >90% in MM cell lines. Primary, simple reciprocal IgH translocations, which are present in both MGUS and MM, involve many partners and provide an early immortalizing event. Four chromosomal partners appear to account for the majority of primary IgH translocations: 11q13 (cyclin D1), 6p21 (cyclin D3), 4p16 (FGFR3 and MMSET), and 16q23 (c-maf). They are mediated primarily by errors in IgH switch recombination and less often by errors in somatic hypermutation, with the former dissociating the intronic and 3' enhancer(s), so that potential oncogenes can be dysregulated on each derivative chromosome (e.g., FGFR3 on der14 and MMSET on der4). Secondary translocations, which sometimes do not involve Ig loci, are more complex, and are not mediated by errors in B cell specific DNA modification mechanisms. They involve other chromosomal partners, notably 8q24 (c-myc), and are associated with tumor progression. Consistent with MM being the malignant counterpart of a long-lived PC, oncogenes dysregulated by primary IgH translocations in MM do not appear to confer an anti-apoptotic effect, but instead increase proliferation and/or inhibit differentiation. The fact that so many different primary transforming events give rise to tumors with the same phenotype suggests that there is only a single fate available for the transformed cell.     

Both IGH translocations and chromosome 13q deletions are early events in monoclonal gammopathy of undetermined significance and do not evolve during transition to multiple myeloma.

Molecular and genetic events associated with the transition from monoclonal gammopathy of undetermined significance (MGUS) to multiple myeloma (MM) are still poorly characterized. We investigated serial bone marrow specimens from 11 patients with MGUS who eventually progressed to MM (MM post-MGUS) by interphase fluorescence in situ hybridization for immunoglobulin heavy-chain gene (IgH) translocations and chromosome 13q deletions (del(13q)). In nine patients, IgH translocations were present both in MGUS and MM post-MGUS plasma cells, including three t(11;14)(q13;q32) and one t(4;14)(p16;q32), which was observed already 92 months prior to MM. Similarly, all five MM patients with del(13q) had this aberration already at the MGUS stage. Two patients without IgH translocation and del(13q) had chromosomal gains suggesting hyperdiploidy, but IgH translocations and/or del(13q) did not emerge at MM post-MGUS. IgH translocations and del(13q) are early genetic events in monoclonal gammopathies, suggesting that additional events are required for the transition from stable MGUS to progressive MM.     

Frequent occurrence of CCND1 deregulation in patients with early stages of plasma cell dyscrasia

Chromosomal translocations involving the immunoglobulin heavy chain gene ( IgH ) and nonrandom protooncogene loci are the hallmark of genetic alterations found not only in multiple myeloma (MM), but also in premalignant stages of MM, including monoclonal gammopathy of undetermined significance (MGUS) and smoldering myeloma (SMM). We studied the frequency of IgH (14q32) rearrangements and their partner chromosomes in 16 Japanese patients with MGUS (13 cases), and SMM (3 cases) by means of interphase double-color fluorescence in situ hybridization (DCFISH) applied to purified plasma cells and using CD138-bead selection. IgH rearrangement was recognized in nine of the patients (56.3%). Protooncogene loci juxtaposed to IgH were identified in seven cases including CCND1 (11q13) in six cases and FGFR3 (4p16) in one. Four out of the six t(11;14)-positive cases showed nuclear staining of the cyclin D1 protein, whereas none of the seven t(11;14)-negative cases did. Moreover, neither MUM1 (6p25)- IgH nor MAFB (20q11)- IgH fusion signals were observed. This suggests to us that cyclin D1 deregulation due to the presence of t(11;14) is involved in the early development of plasma cell neoplasms, and that this event alone is not enough for the development of symptomatic myeloma. (Cancer Sci 2003; 94: 350-354)     

Frequent inactivation of the cyclin-dependent kinase inhibitor p18 by homozygous deletion in multiple myeloma cell lines: ectopic p18 expression inhibits growth and induces apoptosis.

Multiple myeloma (MM) is a clonal neoplasm of plasma cells which offers an excellent model to study multistep molecular oncogenesis. In 20-25% of primary tumors and cell lines examined, cyclin D1 is overexpressed due to the translocation t(11;14)(q13;q32). We have characterized cyclin-dependent kinase inhibitor p15 (CDKN2B), p16 (CDKN2A) and p18 (CDKN2C) deletions in cyclin D1-expressing and non-expressing MM cell lines. p18 was found to be frequently deleted (38%); in some cases p18 deletions coexisted with hemizygous p16 deletion. To examine the function of p18 as a putative tumor suppressor in myeloma cells, a zinc-inducible p18 construct was stably transfected into KMS12, a MM cell line with biallelic p18 and monoallelic p16 deletions as well as cyclin D1 overexpression. Ectopic expression of p18 caused 40-45% growth suppression as determined by trypan blue exclusion and MTS assays. p18 induction also resulted in apoptosis, suggesting that inhibition of the cyclin D1/CDK/pRb pathway in these tumor cells could be a crucial step toward the induction of tumor regression via apoptotic cell death. This cell cycle pathway is thus frequently mutated and provides a potentially novel target for gene therapeutic or pharmacologic approaches to human myeloma.     

Inhibition of cell cycle progression by dual phosphatidylinositol-3-kinase and mTOR blockade in cyclin D2 positive multiple myeloma bearing IgH translocations

Multiple myeloma (MM) is a clinically and genetically heterogenous cancer where tumour cells have dysregulated expression of a D-type cyclin, often in association with a recurrent IgH translocation. Patients whose tumour cells express cyclin D2, with the translocation t(4;14) or t(14;16), generally have more proliferative disease and inferior outcomes. The phosphatidylinositol-3-kinase (PI3K) pathway is a major regulator of D-type cyclin expression and cell cycle entry. We evaluated the effect of PI3K pathway blockade on cell cycle behaviour in MM cells, investigating differences between cyclin D2- and cyclin D1-expressing tumours. MM cell lines and primary bone marrow CD138(+) MM cells were exposed to the pan-PI3K/mTOR inhibitor, PI-103, and assessed for cell cycle profiles, [(3)H]-thymidine uptake and cell cycle proteins. We report, in both cell lines and primary MM cells, that PI-103 induced cell cycle arrest with downregulation of cyclin D2 and CDK4/6 in MM cells expressing cyclin D2 via t(4;14) or t(14;16) translocations. Cells expressing cyclin D1 via t(11;14) were insensitive to PI-103, despite exhibiting inhibition of downstream signalling targets. In primary MM cells, PI-103 enhanced the anti-proliferative effects of anti-MM agents. Treatment paradigms including blockade of the PI3K/mTOR pathway should be targeted at patients with IgH translocations associated with cyclin D2 overexpression.     

Ras oncogene expression and DNA content in plasma cell dyscrasias: a flow cytofluorimetric study

Using bivariate flow cytofluorometry, we have determined the nuclear DNA distribution and the expression of the p21 protein (coded by the Ha-ras oncogene) in the bone marrow (BM) cells of five solid tumour patients having histologically normal BM and in those of 57 patients with plasma cell dyscrasia (28 with monoclonal gammopathies of undertermined significance, MGUS, and 29 with multiple myeloma, MM). All normal and MGUS and 21/29 (72.4%) MM BM had diploid modal DNA content and 8/29 (27.6%) MM BM had both diploid and hyperdiploid cell populations. In normal and MGUS BM, the level of the p21 oncoprotein was low and uniform in all G0/G1, S and G2 cells (median fluorescence values in arbitrary units were 6.1 and 7.5, respectively). The level of p21 was increased both in different aliquots of G0/G1 cells and in the S and G2 cells in diploid MM (median value for G0/G1 cells was 20), and especially in MM with hyperdiploid clones (median value for hyperdiploid cells was 40.5, P less than 0.005 with respect to normal and MGUS BM and less than 0.005 with respect to diploid MM BM). The p21 expression was greater in patients with advanced (stage III) than in earlier MM (stages I + II) (P less than 0.005), and it was directly related to the BMPC infiltration (r = 0.7; P less than 0.005). Since p21 expression is greater in MM than in both normal and MGUS BM, Ha-ras could be involved in the malignant plasma cell transformation that distinguishes MM from MGUS.     

Chromosome analysis of 31 Wilms' tumors

Cytogenetic analysis was done on 31 Wilms' tumors, including 2 renal tumors of clear cell sarcoma type, using short term cultures of primary tumors and/or nude mouse passages. Nonrandom secondary chromosome abnormalities, in particular, were noted as evidence of clonal evolution. Apparently normal karyotypes were found in 5 Wilms' tumors, all in patients less than or equal to 22 months old, and in one clear cell sarcoma. Abnormal karyotypes were seen in 25 tumors (80%); 6 were pseudodiploid, 3 were hypodiploid, and 16 (52%) were hyperdiploid, of which 8 had a modal number of 47-49 and 8 had a modal number of 50-55. Nonrandom structural abnormalities involved 1p/1q, 11p, 7p/7q, 16p/16q, 12q, and 17p/17q. Nonrandom numerical abnormalities included +6, +8, and +18. Trisomy 12 was the most common abnormality, structural or numerical, seen in 52% of tumors (81% of the hyperdiploid). In 2 tumors the +12 was the only apparent abnormality; in 1 other tumor an i(12q) was seen, suggesting that +12 may have special significance in the clonal progression of Wilms' tumor. Informative karyotypes of 68 Wilms' tumors from other reports were reviewed and compared to results in this series.     

Detection of chromosome 13q deletions and IgH translocations in patients with multiple myeloma by FISH: comparison with karyotype analysis

Multiple myeloma (MM) is a plasma cell dyscrasia characterized by frequent 13q deletions and IgH translocations that have clinical prognostic significance. We evaluated clonal plasma cells by interphase fluorescence in situ hybridization (FISH) and combined with immunofluorescence detection of cytoplasmic light chain (cIg-FISH) for the presence of 13q deletions and IgH translocations. The FISH results were compared with conventional cytogenetic analysis. Of the 25 bone marrow specimens from MM patients, 11 (44%) had 13q deletions. IgH translocations involving cyclin D1 (t(11;14)) and FGFR3 (t(4;14)) were found in 32 and 36%, respectively. P53 deletions were detected in 20% of the cases. One patient had coexistence of t(11;14) and t(4;14), which has not been previously reported. Conventional cytogenetic analysis was performed in 15 cases and revealed complex numerical and structural changes in 7. Karyotype analysis failed to detect 3 of 6 cases with 13q deletions, and also missed most of the IgH translocations and p53 deletions detected by cIg-FISH. On the other hand, the complex numerical and structural changes shown by conventional cytogenetics were not demonstrated by interphase FISH. Since 13q deletions, IgH translocations and a hypodiploid karyotype are significant prognostic factors for MM, our study illustrates the importance of combining conventional cytogenetics with interphase FISH analysis in patients with MM.     

Cyclin D1 overexpression is not a specific grouping marker, but may collaborate with CDC37 in myeloma cells

Cyclin D1 is a positive-regulator of the cell cycle and is overexpressed in myeloma cells with t(11;14)(q13;q32). First, we analyzed whether there was a correlation between cyclin D1 overexpression and the presence of Ki67-positive myeloma cells in multiple myeloma (MM). Cyclin D1 overexpression was examined by competitive RT-PCR. Then we found these two markers were present independently in a given case. FISH analysis revealed that cyclin D1 over-expression was caused by t(11;14)(q13;q32) or extra copies of B-cell leukemia/lymphoma-1 (BCL-1/CCND1), and unknown mechanism without them. We compared the gene expression between myeloma cells with cyclin D1 overexpression and those without it using cDNA microarray analysis. Analysis of the expression profiles showed that the significantly up-regulated genes included cyclin D1, cell division cycle 37 (CDC37) and B-cell leukemia/lymphoma-2 (BCL-2), while the down-regulated genes included cyclin D2 and CD9 antigen (p24) in MM cases with cyclin D1 overexpression. However, hierarchical clustering analysis of the data showed that myeloma cells of MM cases with cyclin D1 overexpression could not be distinguished clearly from those without it. Real-time RT-PCR showed that the expression of CDC37 gene was significantly up-regulated in MM patients with cyclin D1 overexpression compared with those without it (p=0.0418). However, there was no significant difference in BCL-2 gene (p=0.5748). These results suggested that MM cases with cyclin D1 overexpression do not constitute a specific group, and cyclin D1 overexpression may not be caused only by abnormality of the BCL-1/CCND1 gene. The CDC37 may collaborate with cyclin D1 in progression of MM.     

Establishment and characterization of a cytogenetically complex Chinese multiple myeloma-derived cell line with homozygous p53 deletion and cyclin E overexpression

We describe the establishment and characterization of a new myeloma-derived cell line (MM17), originating from the sacral plasmacytoma of a 54-year-old Chinese woman diagnosed with multiple myeloma (MM). MM17 was confirmed morphologically and immunophenotypically to be clonal plasma cells positive for CD38 and CD138 and negative for EBV marker. Authenticity was confirmed using comparative genomic hybridization and DNA fingerprinting studies on bone marrow aspirate, sacral tumor tissue and MM17. Combined G-banding and multicolor fluorescence in situ hybridization analyses demonstrated a primarily hypodiploid karyotype with two sidelines sharing common stemline aberrations: +6, -7, -10, -13, -14, -17, -X, der(1;17)(q10;q10), t(2;7)(q23;q11.2), t(8;14)(q24;q32) and ins(16;1)(q13;?q22q41); and a number of hypertriploid cells. The involvement of p53 alteration and cyclin E overexpression, both with relevance to the induction of chromosomal instability, was investigated in MM17 and together with two other MM derived cell lines (U266 and IM-9) for cyclin E expression. Homozygous deletion of p53 gene hitherto not reported in MM, was detected. Both MM17 and U266 with complex cytogenetic aberrations demonstrated overexpression of cyclin E1 and E2, whereas IM-9 with a normal karyotype showed cyclin E2 but not E1 overexpression. These data suggested that E1 but not E2 overexpression was associated with chromosomal abnormalities observed in MM17 and U266, which provides the first supporting evidence for the link of cyclin E and chromosomal instability in MM. This is the first characterized Chinese MM-derived cell line with homozygous p53 deletion which may serve as a valuable in vitro system for studying MM pathogenesis particularly for Chinese.     

14q32 translocations and monosomy 13 observed in monoclonal gammopathy of undetermined significance delineate a multistep process for the oncogenesis of multiple myeloma. Intergroupe Francophone du Myélome.

Clonal plasma cells in monoclonal gammopathy of undetermined significance (MGUS) have been shown to bear copy number chromosome changes. To extend our knowledge of MGUS to structural chromosomal abnormalities, we have performed fluorescence in situ hybridization experiments with probes directed to the 14q32 and 13q14 chromosomal regions in 100 patients with either MGUS or smoldering multiple myeloma (SMM). 14q32 abnormalities were observed in at least 46% of patients with MGUS/SMM, with these abnormalities being present in the majority of clonal plasma cells. Whereas t(11;14)(q13;q32) occurs in 15% of MGUS/SMM patients, an incidence similar to that of overt multiple myeloma (MM) patients, translocation t(4;14)(p16;q32) is observed in only 2% of these cases [P = 0.002 for difference with t(11;14)], as compared with 12% in MM patients (P = 0.013). Monoallelic deletions of the 13q14 region were found in 21% of patients, with two types of situations. In half of the evaluable patients, and especially in patients with SMM, the deletion is present in the majority of clonal plasma cells, as in MM, whereas in the other half of the evaluable patients (essentially in MGUS patients), it is observed in subclones only. These data enable us to elaborate a plasma cell oncogenesis model from MGUS to MM.     

Hyperdiploidy is a common finding in monoclonal gammopathy of undetermined significance and monosomy 13 is restricted to these hyperdiploid patients.

PURPOSE: Two pathways, hyperdiploid and nonhyperdiploid, are proposed for progression to plasma cell neoplasia. Implication of monosomy 13 (Delta13) is unclear in monoclonal gammopathy of undetermined significance (MGUS), and data on DNA content of plasma cells [DNA index (DI)] are rare. EXPERIMENTAL DESIGN: We ascertained DI in 169 multiple myeloma (MM) and 96 MGUS patients. Interphase fluorescence in situ hybridization (FISH) coupled to cytoplasmic staining of specific Ig (cIg-FISH) was done to look for trisomies and to ascertain Delta13. RESULTS: Hyperdiploidy and hypodiploidy were found in 54% and 11.5% of MGUS patients and in 59.5% and 25% of MM patients, respectively. In MGUS patients tested using probes for odd chromosomes, cIg-FISH showed association between trisomies for chromosomes 3, 7, 9, 11, or 15 and hyperdiploidy. Delta13 was found in 45.3% and 24.6% of MM and MGUS patients, respectively. Most Delta13 cases observed in MGUS were found within hyperdiploid clones, 38% versus 11% in hypodiploid cases, in sharp contrast with the occurrence of Delta13 in MM patients, 31.9% and 76.3%, respectively. That peculiar distribution of Delta13 according to DI persisted with other thresholds used to ascertain hyperdiploidy, such as DI >or= 1.05. A strong relationship between IgA peak and hypodiploidy (P = 0.007) was only observed in MM, whereas lambda light chain was significantly associated with hypodiploidy in MGUS (P = 0.001) and MM (P = 0.05). Hyperdiploidy shows similar pattern in MGUS and MM. CONCLUSION: This fits well a hyperdiploid pathway leading to MM after a preceding MGUS stage. Yet-to-be-determined secondary event(s) needs to occur for the transition to MM, unrelated to changes in chromosome number or to loss of chromosome 13. In contrast, the "nonhyperdiploid" pathway needs to be clarified further because hypodiploidy is less common in MGUS than in MM and Delta13 is rare in hypodiploid MGUS patients compared with hypodiploid MM patients.     

Individualizing therapy using molecular markers in multiple myeloma

Multiple myeloma is a heterogeneous disease with complex genetics, including a variety of primary and secondary genetic events that contribute to disease pathogenesis. Good risk genetics include hyperdiploidy (approximately 40%) and cyclin D translocations (t[11;14] and t[6;14]; approximately 18%). Poor risk genetics include t(4;14), approximately 15%; MAF translocations (t[14;16]) and t[14;20]; approximately 8%); secondary genetic events such as deletion p53, and abnormal cytogenetics (deletion 13 or hypodiploidy). Proliferation is the other poor risk feature, as measured by either plasma cell labeling index, beta(2)-microglobulin, or gene expression profiling; it is identified in all genetic subtypes and not yet captured by any genetic marker. Altogether, approximately 75% of patients are good risk and if eligible, do well with high-dose melphalan and autologous stem cell transplantation. In contrast, about 25% of the patients have poor risk features and receive only transient benefit from this approach. We propose the Mayo Stratified Myeloma and Risk-Adapted Therapy. Stem cell transplantation is deferred in patients with high-risk molecular markers, and in all patients, response is followed closely and determines the individual timing and sequence of therapeutic regimens.     

Alterations of the cyclin D1/pRb/p16(INK4A) pathway in multiple myeloma.

The retinoblastoma protein (pRb), p16(INK4A), D-type cyclins, and their partners cyclin-dependent kinase (CDK) 4 and 6 constitute a G(1) regulatory pathway commonly targeted in tumorigenesis. Several malignancies show a reciprocal correlation between genetic alterations of single members of the pRb pathway. Therefore, we determined the frequency of Rb deletions and cyclin D1 alterations by fluorescence in situ hybridization as well as 5' CpG island hypermethylation of the p16(INK4A)gene using methylation-specific polymerase chain reaction in bone marrow mononuclear cells from 82 individuals with plasma cell disorders. Alterations in at least one of the components of the pathway were found in 75%. Cyclin D1 translocations or amplifications were detected in 14/82 (17.1%), Rb deletions at 13q14 in 23/82 (28%) of the cases, including three (3.6%) homozygous deletions. p16(INK4A) was hypermethylated in 33/57 (57.9%) of the samples. Further analysis revealed a highly significant correlation between cyclin D1 alterations and extramedullar or leukemic myeloma manifestations (P = 0.014; Fisher's test). Whereas Rb deletions seemed to occur alternatively to cyclin D1 alterations, no reciprocal correlation was found between p16(INK4A) hypermethylations and cyclin D1 or Rb locus aberrations. Cyclin D1 locus alterations and Rb deletions were associated with a significantly worse prognosis whereas p16(INK4A) hypermethylation had no impact on survival. We conclude that cyclin D1 and Rb aberrations seem to occur as alternative events in plasma cell malignancies and contribute to clinical course and prognosis. In contrast, although p16(INK4A) hypermethylation is frequent, inactivation of p16(INK4A) seems not to be involved in the pathogenesis of plasma cell disorders.     

The prognostic significance of cytogenetics and molecular profiling in multiple myeloma

Multiple myeloma (MM) is a plasma cell malignancy characterized by very complex cytogenetic and molecular genetic aberrations. In newly diagnosed symptomatic patients, the modal chromosome number is usually either hyperdiploid with multiple trisomies or hypodiploid with one of several types of immunoglobulin heavy chain (Ig) translocations. The chromosome ploidy status and Ig rearrangements are two genetic criteria that are used to help stratify patients into prognostic groups based on the findings of conventional cytogenetics and fluorescence in situ hybridization (FISH). In general, the hypodiploid group with t(4;14)(p16;q32) or t(14;16)(q32;q23) is considered a high-risk group, while the hyperdiploid patients with t(11;14)(q13;q32) are considered a better prognostic group. As the disease progresses, it becomes more proliferative and develops a number of secondary chromosome aberrations. These secondary aberrations commonly involve MYC rearrangements, del(13q), del(17p), and the deletion of 1p and/or amplification of 1q. Of the secondary aberrations, del(17p) is consistently associated with poor prognosis. All of these cytogenetic aberrations and many additional ones are now identified by means of high resolution molecular profiling. Gene expression profiling (GEP), array comparative genomic hybridization (aCGH), and single-nucleotide polymorphism (SNP) arrays have been able to identify novel genetic aberration patterns that have previously gone unrecognized. With the integration of data from these profiling techniques, new subclassifications of MM have been proposed which define distinct molecular genetic subgroups. In this review, the findings from conventional cytogenetics, interphase FISH, GEP, aCGH, and SNP profiles are described to provide the conceptual framework for defining the emerging molecular genetic subgroups with prognostic significance.     

Fluorescence in situ hybridization analysis of chromosome aberrations in 60 Chinese patients with multiple myeloma

Conventional cytogenetic analysis is often hampered owing to the low mitotic index of multiple myeloma (MM) cells in bone marrow samples of MM. Interphase fluorescence in situ hybridization (I-FISH) analysis combined with magnetic-activated cell sorting (MACS) has substantially enhanced the sensitivity of cytogenetic analysis. Here, we used I-FISH to explore the incidence of chromosomal abnormalities in 60 Chinese patients with newly diagnosed MM. Five different specific probes for the regions containing 13q14.3 (D13S319), 14q32 (IGHC/IGHV), 1q21, 1p12, and 17p13 were used to detect chromosomal aberrations, and LSI IGH/CCND1, LSI IGH/FGFR3, and LSI IGH/MAF probes were further applied to detect t(11;14)(q13;q32), t(4;14)(p16;q32), and t(14;16)(q32;q23) in patients with 14q32 rearrangement. Fifty of the patients (83.3%) had at least one type of abnormalities regarding the regions analyzed. Nine patients (15%) had one abnormality; 10 patients (16.7%) had two abnormalities; 31 patients (51.7%) had three or more abnormalities. The most frequent abnormality in the patients was illegitimate IgH rearrangement (70%), followed by 13q14 deletion (63.3%), 1q21 amplification (61.7%), 1p12 deletion (33.3%), and 17p13 deletion (13.3%). These aberrations are not randomly distributed, but strongly interconnected. Patients with 17p13 deletion or t(4;14)(p16;q32) had significant higher ??2-microglobulin level (P?<?0.05). However, all these abnormalities had no correlation with age, gender, disease stage, and Ig isotype; yet, it was showed that the frequencies of the individual chromosomal abnormalities were very high. Taken together, MACS in combination with I-FISH may be a promising tool to detect the molecular cytogenetic abnormalities of MM.     

Deletions of chromosome 13q in monoclonal gammopathy of undetermined significance.

Since deletion of chromosome 13q is a clinically relevant feature in multiple myeloma (MM), we analyzed bone marrow plasma cells from 29 patients with monoclonal gammopathy of undetermined significance (MGUS) to investigate the chromosome 13 status in MGUS. Studies were performed by interphase fluorescence in situ hybridization (FISH) with a panel of 13q14-specific probes (RB1, D13S319, D13S25, D13S31). Plasma cells with a deletion of at least one of the 13q14 loci were detected in 13 patients (44.8%) with MGUS. In five patients (17.2%), deletions of all four 13q14-specific probes were observed, and the additional deletion of a 13q telomeric region (D13S327) suggested loss of the entire 13q arm or monosomy 13. Loss of 13q14 was observed to be monoallelic and to occur in 11.0 to 35.0% of plasma cells (cut-off levels for a deletion <10% with all probes). Nine of 17 patients (52.9%) with MM progressing from a pre-existing MGUS had evidence for a deletion of 13q14 as determined by FISH with the RB1 probe. These results suggest that deletion of 13q14 is an early event in the development of monoclonal gammopathies, but its role for the eventual progression to MM remains to be determined prospectively.     

A new multiple myeloma cell line, MEF-1, possesses cyclin D1 overexpression and the p53 mutation

BACKGROUND: The t(11;14)(q13;q32) translocation with cyclin D1 overexpression commonly is found in multiple myeloma (MM) and in mantle cell lymphoma (MCL). Several reports have shown that p53 mutations in MCL lead to blastoid transformation and a worse prognosis; however, the role of p53 mutations in MM with t(11;14) is unclear. METHODS: In this study the authors describe a patient with MM with t(11;14) and a p53 mutation at presentation and characterized a cell line, MEF-1, established from this patient. Immunohistochemical analysis of p53 and cyclin D1 proteins was performed. The p53 gene was analyzed by polymerase chain reaction-single strand conformation polymorphism and direct sequencing. The expression of cyclin D1 mRNA was examined by Northern blot analysis. RESULTS: MEF-1 had t(11;14) with overexpression of cyclin D1 mRNA and produced immunoglobulin kappa-light chain. MEF-1 had a mutation in exon 7 (codon 255-257) of the p53 gene, which was noted in the patient's myeloma cells. CONCLUSIONS: p53 mutations may be important genetic events in disease progression of MM with t(11;14). The MEF-1 cell line may be a useful tool to study mechanisms of progression in MM based on abnormalities of the cyclin D1 gene.