Haematologica, Vol 92, Issue 2, 262-263 doi:10.3324/haematol.10744
Copyright © 2007 by Ferrata Storti Foundation
New types of MYST3-CBP and CBP-MYST3 fusion transcripts in t(8;16)(p11;p13) acute myeloid leukemias
Anne Murati*,°,
José Adélaïde*,
Benoît Quilichini#,
Virginie Rémy*,°,
Danielle Sainty°,
Anne-Marie Stoppa@,,
Philippe Bernard^,
Sylviane Olschwang*,°,
Daniel Birnbaum*,
Max Chaffanet*,
Marie-Joëlle Mozziconacci*,°
* Centre de Recherche en Cancérologie de Marseille, Département d’Oncologie Moléculaire, Institut Paoli-Calmettes et UMR599 Inserm, Marseille, France
° Département de BioPathologie, Institut Paoli-Calmettes, Marseille, France;
# Laboratoire de Cytogénétique, Hôpital de la Timone, Marseille, France
@ Département d’Hématologie, Institut Paoli-Calmettes, Marseille, France
^ Service d’Oncologie, Hôpital Sainte-Anne, Toulon, France
Correspondence: Marie-Joëlle Mozziconacci, Départements d’Oncologie Moléculaire et de BioPathologie, Institut Paoli-Calmettes, 232 Boulevard de Sainte-Marguerite, 13009 Marseille, France. Phone: international +33.4.91223447. Fax: international +33.4.91223544. E-mail: mozziconaccimj{at}marseille.fnclcc.fr
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ABSTRACT
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The t(8;16)(p11;p13) translocation, associated with poor prognosis acute monocytic leukemia, fuses MYST3 on chromosome region 8p11 to CBP on chromosome region 16p13. Two types of MYST3-CBP and CBP-MYST3 fusion transcripts have been identified in patients. We describe two new types of MYST3-CBP transcripts and a new primer set.
Key words: MYST3-CBP, CBP-MYST3, t(8;16), acute myeloid leukemia.
Translocation t(8;16)(p11;p13) is found in 6.5% of acute myeloid leukemias (AML) of the M4/M5 FAB subtype. These poor prognosis AML are associated with erythrophagocytosis in blast cells. The translocation fuses MYST3/MOZ on chromosome region 8p11, which encodes a histone acetyltransferase (HAT), to CBP/CREBBP on chromosome region 16p13, which encodes a transcriptional co-activator and acetyltransferase. 1–4 Reverse transcriptase-polymerase chain reaction (RT-PCR) identified three types of MYST3-CBP and CBP-MYST3 fusion transcripts in some previous studies,1–7 but was unsuccessful in some others suggesting the existence of translocation variants.1,8,9 We describe here two new types of MYST3-CBP fusion transcripts.
Patient #1, a 55-year old female presented with AML-M5a 2 years after breast cancer. The white blood cell (WBC) count was 2.9x109/L with 17% blast cells. The bone marrow aspirate was hypercellular with 90% of monocytic blast cells with features of erythrophagocytosis. The karyotype showed a t(8;16) and a t(11;19) in all mitoses, and an additional 3q deletion and 8q tetrasomy in 65% of mitoses. The patient was treated with idarubicin and cytosine arabinoside, followed by autologous stem cell transplantation. She is still alive in complete remission.
Patient #2, a 72-year old man presented with AML-M5a. The WBC count was 51.3x109/L with 76% blast cells. The bone marrow aspirate was hypercellular with 68% of monocytic blast cells with features of hemophagocytosis. A complex t(8;16) was found in 65% of mitoses with insertion of 8q material between 16p and 8p on the der(16). The patient died 1 month after diagnosis.
On RNA extracted from bone marrow cells of the patients we used nested RT-PCR as described by Schmidt et al.4 for the detection of type I (MYST3 exon 16-CBP exon 3) (Figure 1A, B) and type II (MYST3 exon 16-CBP exon 4) MYST3-CBP fusion transcripts, as well as type I CBP-MYST3 fusion transcript (CBP exon 2-MYST3 exon 17) (Figure 1C, D). PCR products were sequenced after purification.
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Figure 1. RT-PCR and partial sequences of MYST3-CBP and CBP-MYST3 fusions. A–B. Nested PCR with the published4 primer combination exon 16 MYST3_3536F/exon 5 CBP_1201R and exon 16 MYST3_3558F/exon 3 CBP_404R amplifies a ~ 330 bp fragment in the type I control. Cases 1 and 2 are not amplified. C–D. The reciprocal CBP-MYST3 fusion transcript is observed with nested PCR and primer combination exon 2 CBP_96F/exon 17 MYST3_3953R and exon 2 CBP_174F/exon 17 MYST3_3844R. A 229bp fragment and a 943 bp fragment are amplified in the type I control and in case 1, respectively. E–F. PCR with primer combination exon 15 MYST3_3319F/exon 5 CBP_1201R amplifies a ~ 350 pb fragment in case 1 and a 176 bp fragment in case 2, suggesting a different breakpoint location. No fusion is amplified in the type I control. Normal: RNA lymphocytes; Control: RNA from patient with MYST3-CBP and CBP-MYST3 type I transcripts; Genomic: genomic DNA; M: 100 bp DNA molecular weight ladder. The NMA gene was used to control the efficiency of the RT-PCR.
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MYST3-CBP fusion transcripts were not detected in either of the two cases. A 943bp CBP-MYST3 product was obtained in case 1 but not in case 2. A nested PCR with two forward primers in CBP exons 3 and 4 and two reverse primers in MYST3 exons 17 and 16 was also negative in case #2, as expected because of the insertion of 8q material. Sequencing of the 943 bp product showed an in-frame fusion between CBP exon 3 and MYST3 exon 16 whereas the fusion obtained for the type I transcript used as positive control in this reaction showed the expected 229bp fragment4 (Figure 1C, D). To detect MYST3-CBP transcripts we designed a forward primer (Table 1) in MYST3 exon 15 (MYST3_3319F) and the RT-PCR reaction with the CBP exon 5 reverse primer (CBP_1201R) yielded a ~ 350bp for case 1 and a ~ 180bp for case 2. Sequence analysis showed an in-frame fusion between MYST3 exon 15 and CBP exon 4, and between MYST3 exon 15 and CBP exon 5, respectively (Figure 1E, F).
Up to now, 49 AML with t(8;16)(p11;p13) have been reported; 94% of these are monocytic leukemias (mostly of M5 subtype).10 Sixteen cases have been characterized by RT-PCR.1–7 Type I transcript is the most frequent fusion product (14/16 cases); breakpoints map in MYST3 intron 16. Type II is a theoretical, out-of-frame sequence and has not been described in patients. In rare cases (2/16) the breakpoint is within MYST3 exon 17,1,3 which is fused to CBP exon 2 or 4 1,3 defining type III. Our two cases thus define the fourth and fifth types, in which the MYST3-CBP fusions have lost MYST3 exon 16, suggesting that the breakpoint occurs in intron 15. Thus, all the reported breakpoints, including those described here, are located downstream of exon 14. The intact MYST domain in the chimeras may be responsible for aberrant chromatin acetylation due to mistargeting of specific HAT activities and may interfere with gene expression. In type I fusions the breakpoint occurs in CBP intron 2. Our two cases involve exons 4 and 5 (cases 1 and 2, respectively) suggesting that the breakpoints occur in intron 3 or 4. As in type I cases, the MYST3-CBP proteins retain most of the CBP functional domains including the HAT domain.
We have established an RT-PCR assay able to detect MYST3-CBP type IV and V with MYST3 exon 15-CBP exon 4 or 5 fusion. At diagnosis, type I may be screened for first and, if negative, completed by the search for types III, IV and V. An alternate strategy may be to use our set of primers followed by a nested PCR to detect type I or III. Our study may also help better monitoring of minimal residual disease of AML with t(8;16).
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Footnotes
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Funding: this work was supported by Inserm and the Institut Paoli-Calmettes.
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References
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- Borrow J, Stanton VP Jr, Andresen JM, Becher R, Behm FG, Chaganti RS, et al. The translocation t(8;16)(p11;p13) of acute myeloid leukaemia fuses a putative acetyltransferase to the CREB-binding protein. Nat Genet 1996;14:33-41.[CrossRef][ISI][Medline]
- Panagopoulos I, Isaksson M, Lindvall C, Bjorkholm M, Ahlgren T, Fioretos T, et al. RT-PCR analysis of the MOZ-CBP and CBP-MOZ chimeric transcripts in acute myeloid leukemias with t(8;16)(p11;p13). Genes Chromosomes Cancer 2000;28:415-24.[CrossRef][ISI][Medline]
- Panagopoulos I, Fioretos T, Isaksson M, Mitelman F, Johansson B, Theorin N, et al. RT-PCR analysis of acute myeloid leukemia with t(8;16)(p11;p13): identification of a novel MOZ/CBP transcript and absence of CBP/MOZ expression. Genes Chromosomes Cancer 2002;35:372-4.[CrossRef][ISI][Medline]
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- Rozman M, Camos M, Colomer D, Villamor N, Esteve J, Costa D, et al. Type I MOZ/CBP (MYST3/CREBBP) is the most common chimeric transcript in acute myeloid leukemia with t(8;16)(p11;p13) translocation. Genes Chromosomes Cancer 2004;40:140-5.[CrossRef][ISI][Medline]
- Tasaka T, Matsuhashi Y, Uehara E, Tamura T, Kakazu N, Abe T, et al. Secondary acute monocytic leukemia with a translocation t(8;16)(p11;p13): case report and review of the literature. Leuk Lymphoma 2004;45:621-6.[CrossRef][ISI][Medline]
- Bernasconi P, Orlandi E, Cavigliano P, Calatroni S, Boni M, Astori C, et al. Translocation (8;16) in a patient with acute myelomonocytic leukemia, occurring after treatment with fludarabine for a low-grade non-Hodgkin’s lymphoma. Haematologica 2000;85:1087-91.[Abstract/Free Full Text]
- Giles RH, Dauwerse JG, Higgins C, Petrij F, Wessels JW, Beverstock GC, et al. Detection of CBP rearrangements in acute myelogenous leukemia with t(8;16). Leukemia 1997;11:2087-96.[CrossRef][ISI][Medline]
- Troke PJ, Kindle KB, Collins HM, Heery DM. MOZ fusion proteins in acute myeloid leukaemia. Biochem Soc Symp 2006;73:23-39.[Medline]
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