RT-PCR and FISH analysis of acute myeloid leukemia with t(8;16)(p11;p13) and chimeric MOZ and CBP transcripts: breakpoint cluster region and clinical implications

Abstract

The translocation t(8;16)(p11;p13) is associated with acute myeloid leukemia displaying monocytic differentiation (AML FAB M4/5) and fuses the MOZ (also named MYST3) gene (8p11) with the CBP (also named CREBBP) gene (16p13). Detection of the chimeric RNA fusions has proven difficult; only three studies have described successful amplification of the chimeric MOZ-CBP and CBP-MOZ fusions by reverse transcriptasepolymerase chain reaction (RT-PCR). We analyzed four cases of AML M4/5 with t(8;16)(p11;p13) by RT-PCR and fluorescence in situ hybridization (FISH) and characterized the reciprocal RNA fusions from three cases. We cloned both genomic translocation breakpoints from one case by long-range PCR and successfully applied RT-PCR to monitor minimal residual disease (MRD) between clinical complete remission and relapse. In three cases, the genomic breakpoints occurred in MOZ intron 16 and CBP intron 2. In one case, no fusion transcript was detected. The available data suggest clustering of t(8;16)(p11;p13) breakpoints in these introns leading to reciprocal in-frame MOZ exon 16/CBP exon 3 and in-frame CBP exon 2/MOZ exon 17 chimeric transcripts in the majority of cases. The described RT-PCR strategy may be valuable both for the routine detection of the t(8;16)(p11;p13) as well as for monitoring of MRD in this prognostically unfavorable patient group.

AML with translocation t(8;16)(p11;p13) demonstrates unique cytomorphological, cytogenetic, molecular and prognostic features

“…For subsequent molecular analyses these stabilized lysates were frozen and stored at À80 1C. RT-PCR was performed as described by Borrow et al 24 and by Schmidt et al 25 Samples that failed to amplify a MYST3-CREBBP product where further screened according to primers as published by Murati et al 26 The microarray sample preparation assay was performed as previously reported. 4,5,27 Microarray data analysis Gene expression data were processed according to the manufacturer's recommendations.…”

NCOA3, a new fusion partner for MOZ/MYST3 in M5 acute myeloid leukemia

“…The translocation results in the fusion of the MYST3 gene (also known as MOZ) on chromosome region 8p11 to the CREBBP gene (also known as CBP) on chromosome region 16p13. [1][2][3] In the same type of AML, the t(10;16)(q22;p13) fuses MYST3-homolog MYST4 to CREBBP 4 and the t(8;22)(p11;q13) fuses MYST3 to CREBBPhomolog EP300. 5 MYST3, MYST4, CREBBP and EP300 encode transcriptional regulators and acetyltransferases.…”

Detection of MOZ-CBP fusion in acute myeloid leukemia with 8;16 translocation

“…Sequencing analysis revealed that type I transcript represented an in-frame fusion between MOZ exon 16 and CBP exon 3, and type II transcript represented an out-of-frame fusion between MOZ exon 16 and CBP exon 5. 5,7,8 Since type I transcript is from an in-frame fusion and is the most successfully detected fusion product, this type of fusion is most likely the critical event in leukemogenesis. Recently, Schmidt et al 5 described a more sensitive RT-PCR assay for the detection of type I MOZ-CBP fusion transcript.…”

“…Fluorescence in situ hybridization (FISH) is an alternative method for the detection of a specific translocation and for monitoring minimal residual disease. Although a few cosmid clones in the CBP region have been used as FISH probes 3,5 to demonstrate the disruption of the CBP gene, no FISH probe targeting MOZ-CBP fusion is currently available to diagnostic laboratories.…”

“…Fluorescence in situ hybridization (FISH) is an alternative method for the detection of a specific translocation and for monitoring minimal residual disease. Although a few cosmid clones in the CBP region have been used as FISH probes 3,5 to demonstrate the disruption of the CBP gene, no FISH probe targeting MOZ-CBP fusion is currently available to diagnostic laboratories.Taking advantage of the completed human genome project, we searched the National Center for Biotechnology Information (NCBI) database and identified bacteria-derived artificial chromosome (BAC) clones in the MOZ and CBP regions, respectively. BAC clones with an average insert size of 70-100 kb are ideal material to be used as FISH probes.…”