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This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Hayman, S. R. Articles by Fonseca, R. Search for Related Content PubMed PubMed Citation Articles by Hayman, S. R. Articles by Fonseca, R. Related Collections Neoplasia Brief Reports Social Bookmarking                   What's this?  Previous Article  |  Table of Contents  |  Next Article  Blood, 1 October 2001, Vol. 98, No. 7, pp. 2266-2268 BRIEF REPORT Translocations involving the immunoglobulin heavy-chain locus are possible early genetic events in patients with primary systemic amyloidosis Suzanne R. Hayman, Richard J. Bailey, Syed M. Jalal, Gregory J. Ahmann, Angela Dispenzieri, Morie A. Gertz, Philip R. Greipp, Robert A. Kyle, Martha Q. Lacy, S. Vincent Rajkumar, Thomas E. Witzig, John A. Lust, and Rafael Fonseca From the Departments of Hematology and Internal Medicine and of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN.     Abstract Top Abstract Introduction Study design Results and discussion References Primary systemic amyloidosis (AL) is a plasma cell (PC) dyscrasia with clinical similarities to multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS), but its molecular basis is poorly understood. Translocations at the immunoglobulin heavy-chain (IgH) locus, 14q32, are likely early genetic events in both MM and MGUS and involve several nonrandom, recurrent, partner chromosomes such as 11q13, 16q23, and 4p16.3. Given the similarities between MM, MGUS, and AL, bone marrow clonal PCs were evaluated in 29 patients with AL using interphase fluorescence in situ hybridization (FISH) combined with immunofluorescence detection of the cytoplasmic light-chain (cIg-FISH) for the presence of 14q32 translocations and the t(11;14)(q13;q32). Of 29 patients studied, 21 (72.4%) showed results compatible with the presence of a 14q32 translocation, and 16 (76.2%) of those had translocation (11;14)(q13;q32) for an overall prevalence of the abnormality of 55%. IgH translocations are common in AL, especially the t(11;14)(q13;q32). (Blood. 2001;98:2266-2268) © 2001 by The American Society of Hematology.     Introduction Top Abstract Introduction Study design Results and discussion References Amyloidosis (AL) is a plasma cell (PC) dyscrasia, similar to multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS),1,2 characterized by the presence of clonal PCs in the bone marrow (BM) and extracellular deposition of amyloidogenic light chains with resultant organ dysfunction and an overall median survival of 1.5 years.1,2 The usually fatal outcome of the disease does not permit long-term observations of clonal growth. Chromosomal translocations involving the immunoglobulin heavy-chain (IgH) locus may be initiating events in the development of several B-cell malignancies,3,4 including MGUS and MM.5 IgH translocations in MM involve a number of recurrent, nonrandom partner chromosomes including 11q13 and 4p16.3.14-16 Using the cytoplasmic light-chain (cIg)-fluorescence in situ hybridization (FISH) technique, we examined 29 patients with AL for the presence of IgH translocations, including t(11;14)(q13;q32).     Study design Top Abstract Introduction Study design Results and discussion References Patient samples Twenty-nine patients (10 women, 19 men; median age, 62 years) were studied. Institutional review board approval was obtained, and all patients gave written, informed consent for a research BM sample to be collected at the time of routine clinical procurement. Diagnostic criteria for AL included histologic proof of amyloid deposition in addition to the presence of a monoclonal protein or demonstration of monoclonal PCs. Five of the 29 patients had previously been treated with alkylating agents. None of the patients had clinical evidence of MM. Cell enrichment and cIg-FISH PCs were selected with CD 138 magnetic microbeads (Miltenyi Biotec, Sunnyvale, CA) after enrichment for mononuclear cells through centrifugation on a Ficoll-Hypaque gradient, density 1.076 (Pharmacia-Biotech, Uppsala, Sweden). Cytospin preparations were prepared according to a slight modification of our previously published cIg-FISH technique.6 We scored 100 PCs in all but 2 cases. One repeatedly failed to hybridize; in the other, we could only score 50 cells for t(11;14)(q13;q32), respectively. Each sample was scored by 2 independent examiners who had a high degree of agreement (r2 = 0.795). FISH probes Break-apart strategy. Probes that hybridize to the variable (VH; cosmid clone) and constant regions (CH; BAC clone) of the IgH locus7 were directly labeled with SpectrumRed and SpectrumGreen, respectively (Vysis, Downers Grove, IL). The presence of a 14q32 translocation was demonstrated by segregation of the signals (break-apart strategy; Figure 1). A normal pattern consisted of 2 pairs of closely associated red and green signals. An abnormal pattern was defined as a distance greater than 3 signal widths between probe pairs. An equivocal pattern was one in which there was loss of a probe signal (1 red-green pair/1 red or 1 red-green pair/1 green), representing deletion, an unbalanced translocation, or technical inability to discern the presence of a signal because of small size or background staining. View larger version (62K): [in this window] [in a new window]   Figure 1. Normal and abnormal patterns of hybridization with both strategies. (A) Hybridization of the VH and CH probes to the correct location at band 14q32 on normal human male metaphases. (B) Normal pattern for the VHCH probes with 2 pairs of closely associated signals. Note the intense blue fluorescence of the cytoplasmic light chain. (C) Cellular signal pattern indicative of a 14q32 translocation. The segregation of a pair of signals (break-apart) in a large percentage of cells allows for the identification of a patient with an IgH translocation. (D) Hybridization of the 14q32 and 11q13 probes to the correct locations on chromosomes 14 and 11, respectively, on normal human male metaphases. (E) Normal pattern for the 14q32/11q13 fusion strategy. There are no fusion signals observed, and there are 2 pairs of nonassociated green (14q32) and red (11q13) signals. (F) Abnormal comigration (fusion) of signals indicative of a t(11;14)(q13;q32). Because of the small signal size arising from the VH portion of the IgH probe (owing to the V/D/J recombination and DNA excision), the pattern most frequently observed is presumed to be that of fusions in the der14 and less commonly in the der11. Fusion strategy. For the detection of the t(11;14)(q13;q32), on separate experiments, we used the same IgH probes (labeled with SpectrumGreen) and a pool of cosmid and P1 clones encompassing all reported breakpoints in human MM cell lines with the t(11;14)(q13;q32)8-10 (labeled with SpectrumRed). The normal pattern was that of 2 pairs of nonassociated signals (2 red/2 green). Fusion of 2 signals in a cell was considered indicative of the translocation. Statistical considerations The normal range of PC patterns was documented previously using cIg-negative cells derived from patient samples and polyclonal PCs from BM donors. A patient was said to have an IgH translocation if the percentage of PCs exceeded the mean ± 3 SD.11 The upper limit of normal for the break-apart and fusion strategies was less than 10% for both. We arbitrarily chose an upper limit of normal of 25% for greater specificity.     Results and discussion Top Abstract Introduction Study design Results and discussion References Break-apart strategy (VH/CH probes) Of these 29 patients, 16 (55%) had definitive evidence of an IgH translocation using the VH/CH strategy (Table 1). The median percentage of abnormal PCs (MAPC) was 55% (range, 27%-89%). Additionally, 5 (17%) patients had an equivocal pattern compatible with a possible IgH translocation (MAPC, 66%; range, 64%-83%). In total, 21 of 29 (72%) patients had evidence of an IgH translocation by the VH/CH probe patterns.                                View this table: [in this window] [in a new window]   Table 1. Patient clinical and laboratory features and associated FISH results t(11;14)(q13;q32) Twelve of 16 (75%) patients with definitive IgH translocations by the VH/CH strategy had a t(11;14)(q13;q32) (MAPC, 87.5%; range, 68%-100%). In addition, 4 of the 5 (80%) patients with an equivocal pattern using the VH/CH strategy also had a t(11;14)(q13;q32) (MAPC, 76%; range, 68%-96%). Altogether, 16 of the 29 (55%) patients had evidence of a t(11;14)(q13;q32), accounting for 76% of all IgH translocations detected (16 of 21 patients). There was no statistically significant difference in the BM percentage PC between patients with and without the abnormality detected by either strategy (Wilcoxon signed-rank test, P > .2) Other Four patients had definitive evidence of an IgH translocation by the break-apart strategy that did not involve the 11q13 locus (MAPC, 64%; range, 44%-91%). One patient had a possible IgH translocation with an equivocal pattern by the VH/CH in 90% of PCs (not involving 11q13). Eight patients did not have evidence of IgH translocations, and 2 of those patients had only 1 pair of VH/CH probes in 55% and 76% of their cells, respectively, reflecting monosomy 14 or deletion of 14q32. We have shown here that most patients with AL harbor IgH translocations in clonal PCs. In AL, the deposition of monoclonal light chains is the cause of the sickness and death associated with the disease, whereas clonal progression is usually not observed.12 Our results support the hypothesis that AL is the same disease entity as MGUS/SMM (at the clonal level), with the clinical differences (phenotype) arising from the amyloidogenic potential of the monoclonal light chain. Most patients with AL have a low degree of BM involvement and rarely have overt MM.13 We speculate the occasional patient with AL (approximately 5%), who also has coexistent MM, may have a lesser amyloidogenic variant of the light chain or a more rapid clonal progression.14 The frequency of t(11;14)(q13;q32) is much greater in AL than has been noted for MM15 or MGUS16-18 in some series but not in others.19 Several facts allow us to conclude that the prevalence of the t(11;14)(q13;q32) we report in AL is correct. We used 2 independent scorers, and both concurred in the scoring of abnormal PCs in each sample with a high degree of agreement. The percentages of abnormal PCs by the break-apart and fusion strategies were similar in each patient, and the abnormalities were usually seen in most of the clonal PC. Finally, we also observed a higher proportion of t(11;14)(q13;q32) in patients with MGUS than in patients with MM.19 The frequencies of other partner chromosomes are being studied to assess the pathogenetic potential of these abnormalities, and we speculate that other partner chromosomes will also be involved.20-22 The results of our study identify cytogenetic lesions in the clonal PCs of patients with AL, similar to those observed in MGUS and MM. In addition, previous work has shown that VL gene sequences are highly somatically mutated, suggesting commonality among these dyscrasias.23 We have previously shown that aneuploidy is equally present in AL, MGUS, smoldering MM, and MM.24 Hallek3 proposes a step-wise transformation model for the development of MM from MGUS, and we propose the inclusion of AL in the model where AL is similar to MGUS and smoldering MM, albeit with an "unlucky" protein.     Footnotes Submitted January 8, 2001; accepted May 17, 2001. Supported in part by Public Health Service grant R01 CA83724-01 from the National Cancer Institute. P.R.G. is supported in part by research grant P01 CA62242 from the National Cancer Institute and by ECOG grant CA21115-25C from the National Cancer Institute. R.F. is a Leukemia and Lymphoma Society Translational Research Awardee. R.F. and P.R.G. are also supported by the Mayo Foundation and the CI-5 Cancer Research Fund, Lilly Clinical Investigator Award of the Damon Runyon-Walter Winchell Foundation. The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734. Reprints: Rafael Fonseca, Department of Hematology and Internal Medicine, 200 First St SW, Bldg W10B, Mayo Clinic, Rochester, MN 55905; e-mail: fonseca.rafael{at}mayo.edu.     References Top Abstract Introduction Study design Results and discussion References 1. Gertz MA, Lacy MQ, Dispenzieri A. Amyloidosis: recognition, confirmation, prognosis, and therapy [review]. Mayo Clin Proc. 1999;74:490-494[Abstract]. 2. Kyle RA, Greipp PR. Amyloidosis (AL): Clinical and laboratory features in 229 cases [review]. Mayo Clin Proc. 1983;58:665-683[Medline] [Order article via Infotrieve]. 3. Hallek M, Leif Bergsagel P, Anderson KC. Multiple myeloma: increasing evidence for a multistep transformation process [review]. Blood. 1998;91:3-21[Free Full Text]. 4. Willis TG, Dyer MJS. The role of immunoglobulin translocations in the pathogenesis of B-cell malignancies. Blood. 2000;96:808-822[Free Full Text]. 5. Bergsagel PL, Chesi M, Nardini E, Brents LA, Kirby SL, Kuehl WM. Promiscuous translocations into immunoglobulin heavy chain switch regions in multiple myeloma. Proc Natl Acad Sci U S A. 1996;93:13931-13936[Abstract/Free Full Text]. 6. Ahmann GJ, Jalal SM, Juneau AL, et al. A novel three-color, clone-specific fluorescence in situ hybridization procedure for monoclonal gammopathies. Cancer Genet Cytogenet. 1998;101:7-11[CrossRef][Medline] [Order article via Infotrieve]. 7. Gabrea A, Bergsagel PL, Chesi M, Shou Y, Kuehl WM. Insertion of excised IgH switch sequences causes overexpression of cyclin D1 in a myeloma tumor cell. Mol Cell. 1999;3:119-123[CrossRef][Medline] [Order article via Infotrieve]. 8. Janssen JW, Vaandrager JW, Heuser T, et al. Concurrent activation of a novel putative transforming gene, myeov, and cyclin D1 in a subset of multiple myeloma cell lines with t(11;14)(q13;q32). Blood. 2000;95:2691-2698[Abstract/Free Full Text]. 9. Vaandrager JW, Schuuring E, Zwikstra E, et al. Direct visualization of dispersed 11q13 chromosomal translocations in mantle cell lymphoma by multicolor DNA fiber fluorescence in situ hybridization. Blood. 1996;88:1177-1182[Abstract/Free Full Text]. 10. Vaandrager JW, Kluin P, Schuuring E. The t(11;14) (q13;q32) in multiple myeloma cell line KMS12 has its 11q13 breakpoint 330 kb centromeric from the cyclin D1 gene [letter]. Blood. 1997;89:349-350[Free Full Text]. 11. Anscombe F. The transformation of Poisson, binomial and negative-binomial data. Biometrika. 1949;35:246-254. 12. Gertz MA, Kyle RA. Amyloidosis: prognosis and treatment [review]. Semin Arthritis Rheum. 1994;24:124-138[CrossRef][Medline] [Order article via Infotrieve]. 13. Gertz MA, Lacy MQ, Dispenzieri A. Amyloidosis: recognition, confirmation, prognosis, and therapy. Mayo Clin Proc. 1999;74:490-494. 14. Kyle RA, Gertz MA. Primary systemic amyloidosis: clinical and laboratory features in 474 cases. Semin Hematol. 1995;32:45-59[Medline] [Order article via Infotrieve]. 15. Fonseca R, Harrington D, Oken MM, et al. Prognostic significance of 13q- deletions and the t(11;14)(q13;q32) in myeloma (MM): an interphase FISH study of 351 patients entered into the Eastern Cooperative Oncology Group E9487 Clinical Trial [abstract]. Blood. 2000;96:547. 16. Avet-Loiseau H, Brigaudeau C, Morineau N, et al. High incidence of cryptic translocations involving the Ig heavy chain gene in multiple myeloma, as shown by fluorescence in situ hybridization. Genes Chromosomes Cancer. 1999;24:9-15[CrossRef][Medline] [Order article via Infotrieve]. 17. Avet-Loiseau H, Li JY, Facon T, et al. High incidence of translocations t(11;14)(q13;q32) and t(4;14)(p16;q32). Cancer Res. 1998;58:5640-5645[Abstract/Free Full Text]. 18. Avet-Loiseau H, Facon T, Daviet A, et al. 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 Myelome. Cancer Res. 1999;59:4546-4550[Abstract/Free Full Text]. 19. Fonseca R, Bailey R, Ahmann G, et al. Translocations involving 14q32 are common in patients with the monoclonal gammopathy of undetermined significance. Proc Int Myeloma Workshop; Stockholm, Sweden:; 1999. 20. Chesi M, Bergsagel PL, Shonukan OO, et al. Frequent dysregulation of the c-maf proto-oncogene at 16q23 by translocation to an Ig locus in multiple myeloma. Blood. 1998;91:4457-4463[Abstract/Free Full Text]. 21. Chesi M, Nardini E, Brents LA, et al. Frequent translocation t(4;14)(p16.3;q32.3) in multiple myeloma is associated with increased expression and activating mutations of fibroblast growth factor receptor 3. Nat Genet. 1997;16:260-264[CrossRef][Medline] [Order article via Infotrieve]. 22. Chesi M, Nardini E, Lim R, Smith K, Kuehl W, Bergsagel P. The t(4;14) translocation in myeloma dysregulates both FGFR3 and a novel gene, MMSET, resulting in IgH/MMSET hybrid transcripts. Blood. 1998;92:3025-3034[Abstract/Free Full Text]. 23. Perfetti V, Ubbiali P, Vignarelli MC, et al. Evidence that amyloidogenic light chains undergo antigen-driven selection. Blood. 1998;91:2948-2954[Abstract/Free Full Text]. 24. Fonseca R, Ahmann GJ, Jalal SM, et al. Chromosomal abnormalities in systemic amyloidosis. Br J Haematol. 1998;103:704-710[CrossRef][Medline] [Order article via Infotrieve]. © 2001 by The American Society of Hematology.   CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this? This article has been cited by other articles: M Deshmukh, K Elderfield, A Rahemtulla, and K N Naresh Immunophenotype of neoplastic plasma cells in AL amyloidosis J. Clin. Pathol., August 1, 2009; 62(8): 724 - 730. [Abstract] [Full Text] [PDF] A. H. Bryce, R. P. Ketterling, M. A. Gertz, M. Lacy, R. A. Knudson, S. Zeldenrust, S. Kumar, S. Hayman, F. Buadi, R. A. Kyle, et al. Translocation t(11;14) and survival of patients with light chain (AL) amyloidosis Haematologica, March 1, 2009; 94(3): 380 - 386. [Abstract] [Full Text] [PDF] M. A. Gertz, M. Q. Lacy, J. A. Lust, P. R. Greipp, T. E. Witzig, and R. A. Kyle Long-term risk of myelodysplasia in melphalan-treated patients with immunoglobulin light-chain amyloidosis Haematologica, September 1, 2008; 93(9): 1402 - 1406. [Abstract] [Full Text] [PDF] T. Bochtler, U. Hegenbart, F. W. Cremer, C. Heiss, A. Benner, D. Hose, M. Moos, J. Bila, C. R. Bartram, A. D. Ho, et al. Evaluation of the cytogenetic aberration pattern in amyloid light chain amyloidosis as compared with monoclonal gammopathy of undetermined significance reveals common pathways of karyotypic instability Blood, May 1, 2008; 111(9): 4700 - 4705. [Abstract] [Full Text] [PDF] N. Gonzalez-Paz, W. J. Chng, R. F. McClure, E. Blood, M. M. Oken, B. V. Ness, C. D. James, P. J. Kurtin, K. Henderson, G. J. Ahmann, et al. 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[Abstract] [Full Text] [PDF] This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Hayman, S. R. Articles by Fonseca, R. Search for Related Content PubMed PubMed Citation Articles by Hayman, S. R. Articles by Fonseca, R. Related Collections Neoplasia Brief Reports Social Bookmarking                   What's this?

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