Ordering of probes surrounding the Ewing&rsquo;s sarcoma breakpoint on chromosome 22 using fluorescent in situ hybridization to interphase nuclei

Abstract Chromosomal translocations involving the immunoglobulin heavy chain (IGH) locus at chromosome 14q32 represent a common mechanism of oncogene activation in lymphoid malignancies. In multiple myeloma (MM), variable chromosome partners have been identified by conventional cytogenetics, including the 11q13, 8q24, 18q21, and 6p21 loci. We and others have recently reported a novel, karyotypically undetectable chromosomal translocation t(4;14)(p16.3;q32) in MM-derived cell lines, as well as in primary tumors. The 4p16.3 breakpoints are relatively scattered and located less than 100 kb centromeric of the fibroblast growth factor receptor 3 (FGFR3) gene or within the recently identified WHSC1 gene, both of which are apparently deregulated by the translocation. To assess the frequency of the t(4;14)(p16.3;q32) translocation in MM, we performed a double-color fluorescent in situ hybridization (FISH) analysis of interphase nuclei with differently labeled probes specific for the IGH locus (a pool of plasmid clones specific for the IGH constant regions) or 4p16.3 (yeast artificial chromosome (YAC) 764-H1 spanning the region involved in breakpoints). Thirty MM patients, the MM-derived cell lines KMS-11 and OPM2, and six normal controls were examined. The identification of a t(4;14) translocation, evaluated as the presence of a der(14) chromosome, was based on the colocalization of signals specific for the two probes; a cutoff value of 15% (mean + 3 standard deviation [SD]) derived from the interphase FISH of the normal controls (range, 5% to 11%; mean ± SD, 8.16 ± 2.2) was used for the quantification analysis. In interphase FISH, five patients (one in clinical stage I, two in stage II, one in stage III, and a plasma cell leukemia) were found to be positive (≈15%). FISH metaphases with split or colocalized signals were detected in only two of the translocated cases and confirmed the pattern found in the interphase nuclei. Furthermore, in three of the five cases with the translocation, FISH analysis with the IGH joining probe (JH) showed the presence of the reciprocal product of the translocation [der(4) chromosome]. Overall, our study indicates that the t(4;14)(p16.3;q32) chromosomal translocation is a recurrent event in MM tumors and may contribute towards the detection of this lesion and our understanding of its pathogenetic and clinical implications in MM.

Download Full-text

Detection of t(4;14)(p16.3;q32) Chromosomal Translocation in Multiple Myeloma by Double-Color Fluorescent In Situ Hybridization

Blood ◽

10.1182/blood.v94.2.724.414k05_724_732 ◽

1999 ◽

Vol 94 (2) ◽

pp. 724-732 ◽

Cited By ~ 2

Author(s):

Palma Finelli ◽

Sonia Fabris ◽

Savina Zagano ◽

Luca Baldini ◽

Daniela Intini ◽

...

Keyword(s):

Multiple Myeloma ◽

In Situ Hybridization ◽

Cell Lines ◽

Fluorescent In Situ Hybridization ◽

Chromosomal Translocation ◽

Fish Analysis ◽

Interphase Nuclei ◽

Igh Locus ◽

Normal Controls

Chromosomal translocations involving the immunoglobulin heavy chain (IGH) locus at chromosome 14q32 represent a common mechanism of oncogene activation in lymphoid malignancies. In multiple myeloma (MM), variable chromosome partners have been identified by conventional cytogenetics, including the 11q13, 8q24, 18q21, and 6p21 loci. We and others have recently reported a novel, karyotypically undetectable chromosomal translocation t(4;14)(p16.3;q32) in MM-derived cell lines, as well as in primary tumors. The 4p16.3 breakpoints are relatively scattered and located less than 100 kb centromeric of the fibroblast growth factor receptor 3 (FGFR3) gene or within the recently identified WHSC1 gene, both of which are apparently deregulated by the translocation. To assess the frequency of the t(4;14)(p16.3;q32) translocation in MM, we performed a double-color fluorescent in situ hybridization (FISH) analysis of interphase nuclei with differently labeled probes specific for the IGH locus (a pool of plasmid clones specific for the IGH constant regions) or 4p16.3 (yeast artificial chromosome (YAC) 764-H1 spanning the region involved in breakpoints). Thirty MM patients, the MM-derived cell lines KMS-11 and OPM2, and six normal controls were examined. The identification of a t(4;14) translocation, evaluated as the presence of a der(14) chromosome, was based on the colocalization of signals specific for the two probes; a cutoff value of 15% (mean + 3 standard deviation [SD]) derived from the interphase FISH of the normal controls (range, 5% to 11%; mean ± SD, 8.16 ± 2.2) was used for the quantification analysis. In interphase FISH, five patients (one in clinical stage I, two in stage II, one in stage III, and a plasma cell leukemia) were found to be positive (≈15%). FISH metaphases with split or colocalized signals were detected in only two of the translocated cases and confirmed the pattern found in the interphase nuclei. Furthermore, in three of the five cases with the translocation, FISH analysis with the IGH joining probe (JH) showed the presence of the reciprocal product of the translocation [der(4) chromosome]. Overall, our study indicates that the t(4;14)(p16.3;q32) chromosomal translocation is a recurrent event in MM tumors and may contribute towards the detection of this lesion and our understanding of its pathogenetic and clinical implications in MM.

Download Full-text

Perilobar Nephrogenic Rests and Chromosome 22

Pediatric and Developmental Pathology ◽

10.2350/10-01-0781-cr.1 ◽

2011 ◽

Vol 14 (6) ◽

pp. 485-492 ◽

Cited By ~ 3

Author(s):

Reena MdZIN ◽

Marianne Phillips ◽

Caitlin Edwards ◽

Ashleigh Murch ◽

Adrian Charles

Keyword(s):

In Situ Hybridization ◽

Fluorescent In Situ Hybridization ◽

Wilms Tumor ◽

Microsatellite Analysis ◽

Chromosome 22 ◽

Hybridization Probe ◽

Genetic Changes ◽

Preferential Loss ◽

Nephrogenic Rests

Perilobar nephrogenic rests (NR) are precursor lesions that may display genetic changes similar to their associated Wilms tumor (WT). Two patients presented with WT, both with perilobar NR and 1 with bilateral, multifocal metachronous WT. Both patients' WT displayed monosomy 22 as the predominant cytogenetic change, and the constitutional cytogenetic analysis was normal. The purpose of our study was to identify at what stage in the morphologic progression from NR to WT the monosomy 22 occurred by using a fluorescent in situ hybridization probe for chromosome 22 in the subtypes of perilobar NR and WT present in both cases. Section and core fluorescent in situ hybridization with a chromosome 22 probe was performed on formalin-fixed, paraffin-embedded tissues containing WT and perilobar NR. We also performed fluorescent-based microsatellite analysis on some of the WT in the bilateral case to determine whether there was a preferential loss of the same allele of chromosome 22. Sclerotic and dormant perilobar NR showed a rate of monosomy 22 of only approximately 30%, but this increased to approximately 50% in hyperplastic and adenomatous NR. Monosomy 22 was present in 60%-80% of nuclei in WT. Microsatellite analysis showed loss of homozygosity, with preferential loss of the same allele of chromosome 22 in the tumors examined. There are differences in the rate of detection of monosomy 22 in perilobar NR and WT, suggesting loss of chromosome 22 in the progression of perilobar NR to WT in a subset of tumors.

Download Full-text