Characterization of an inversion duplication of the short arm of chromosome 8 by fluorescent in situ hybridization

A de novo chromosome aberration in a woman with severe mental retardation and minor anomalies has been characterized cytogenetically. The patient's karyotype was described as 46, XX, inv dup (8)(p12 → p23.1). Previous Southern blot dosage studies with the marker locus D8S7 demonstrated that the patient was monosomic for this locus, suggesting that the rearrangement generated a duplication-deficiency chromosome. We have reinvestigated this patient using fluorescent in situ hybridization with chromosome 8 cosmids and an Alu-PCR product specific for 8p. These studies have confirmed directly that the duplicated chromosome also has undergone deletion. © 1992 Wiley-Liss, Inc.     

Characterization of an inversion duplication of the short arm of chromosome 8 by fluorescent in situ hybridization.

A de novo chromosome aberration in a woman with severe mental retardation and minor anomalies has been characterized cytogenetically. The patient's karyotype was described as 46, XX, inv dup (8)(p12-->p23.1). Previous Southern blot dosage studies with the marker locus D8S7 demonstrated that the patient was monosomic for this locus, suggesting that the rearrangement generated a duplication-deficiency chromosome. We have reinvestigated this patient using fluorescent in situ hybridization with chromosome 8 cosmids and an Alu-PCR product specific for 8p. These studies have confirmed directly that the duplicated chromosome also has undergone deletion.     

Mapping of translocation breakpoints on the short arm of chromosome 19 in acute leukemias by in situ hybridization

Non-random translocation involving the short arm of chromosome 19 are frequently observed in acute leukemias. Recent studies have shown that the 19p13 genes E2A and LYLl, both of which encode helix-loop-helix proteins, lie at two different translocation breakpoints in acute lymphoblastic leukemias (ALL). The E2A gene is involved by the t(1;19)(q23;p13) in acute pre-B-cell leukemias and the LYL1 gene is structurally altered by a t(7;19)(q34;p13) in T-cell ALL. To assess the role of these genes in other leukemia-associated translocations we mapped their locations with respect to the t(11;19)(q23;p13) and t(4;19)(q21;p13) translocation breakpoints carried by T-ALL cell lines SUP-T13 and SUP-T8a, respectively. In situ hybridization studies indicated that the E2A and LYL1 genes are physically distinct from the t(4;19) and t(11;19) breakpoints. Using these and other 19p13 translocation breakpoints as landmarks, we established a partial physical map of 19p: 19pter-E2A-INSR-LYL1-[t(4;19)]-19cen. These data should help guide molecular studies to further characterize 19p13 breakpoints and mapping of genes in this chromosomal region.     

Prenatal detection of de novo duplication of the short arm of chromosome 18 confirmed by fluorescence in situ hybridization (FISH)

We present a patient with developmental delay, minor anomalies, and duplication 18p confirmed by fluorescence in situ hybridization with whole chromosome 18 painting probe (Oncor p5218). Our observation confirms the findings of other investigators that duplication 18p is not associated with major malformations. Am. J. Med. Genet. 80:487–490, 1998. © 1998 Wiley-Liss, Inc.     

In situ hybridization analysis of isodicentric X-chromosomes with short arm fusion

We present here an alternative approach to the study of mosaic cell lines containing dicentric chromosomes. The approach is based on chromosome-specific non-radioactive in situ hybridization with centromere (alpha satellite DNA) probes. The hybridization analysis may be used as an alternative to the C-band analysis, while at the same time to some extent replacing the Q-band analysis as well. The advantage of using in situ hybridization is mainly that it allows the very fast screening of a large number of metaphases. We illustrate this new application of the technique by using it for the analysis of two cases of isodicentric X-chromosomes. The approach is expected to be generally applicable, so that it may be applied to the scoring of other types of chromosomal mosaicism as well.     

Fluorescence in situ hybridization analysis of allelic losses involving the long arm of chromosome 17 in NF1-associated neurofibromas

Neurofibromatosis type 1 (NF1) is a common autosomal dominant condition associated with germline mutations of the NF1 gene located at chromosome band 17q11.2. Molecular analysis of a number of NF1-specific tumors has shown the inactivation of both NF1 alleles during tumorigenesis, supporting the tumor suppressor hypothesis for the NF1 gene. Using interphase dual-color fluorescence in situ hybridization (FISH) technique on paraffin-embedded tissues, we studied 11 plexiform, 4 cutaneous, and 6 subcutaneous neurofibromas. Cytogenetic analysis was conducted using two probes, one specific for the NF1 region (RP11-229K15) and one for the centromeric region of chromosome 17 as control. No large somatic deletions were found. Only in one of the plexiform neurofibromas loss of a whole chromosome 17 was observed. If we assume that dual-color FISH analysis is sensitive enough to detect the majority of large somatic deletions present, then other mutational mechanisms affecting the NF1 gene are probably involved in neurofibroma formation, and other tumor suppressor genes may play an important role in NF1 tumorigenesis.     

Chromosomal localization of a single copy gene by in situ hybridization - human β globin genes on the short arm of chromosome 11

1. The localization of the β globin genes by in situ hybridization to fixed chromosomes is described.
2. The probe used was a [³H]cRN A copy of a genomic clone containing in total 4-4 kb of DNA and including the β globin gene.
3. The evidence for the localization of the gene comes from three pieces of data, ( a ) Chromosome 11 is labelled to double the extent expected if the grains were randomly distributed, ( b ) the extra grains above background are clustered on the short arm of 11 close to the centromere, and ( c ) the absolute number of grains observed is very close to that predicted for a probe of that length by comparison with ribosomal genes. The localization is in agreement with that obtained by other methods.
4. This method could be extended to any gene for which a genomic clone containing at least 5 kb of single copy DNA is available.     

High-resolution mapping of amplicons of the short arm of chromosome 1 in two neuroblastoma tumors by microarray-based comparative genomic hybridization

Deletion of chromosome arm 1p is one of the most frequent genetic alterations in neuroblastoma. However, using conventional comparative genomic hybridization, we have observed amplifications on 1p in 2 neuroblastoma tumors at bands 1p34.2 and 1p36.3, respectively. Using a medium-resolution genomic array containing 178 PACs/BACs from 1p and then 2 high-resolution arrays containing contigs of overlapping PACs/BACs from the amplified regions, we could precisely map and delineate both amplicons. The 1p34.2 amplicon appeared as a homogeneous amplification unit, whereas the 1p36.3 amplicon had a more complex structure, with 2 noncontiguous, highly amplified regions and several moderate amplification units. In this case, fluorescence in situ hybridization analysis confirmed the amplification of several clones and indicated that the 2 highest amplification units corresponded to 2 populations of double minute chromosomes, one of which also contained the MYCN locus. This is the first report of 1p amplifications in primary neuroblastomas.     

Fluorescence in situ hybridization analyses of chromosome band 1p36 in neuroblastoma detect two classes of alterations

Chromosomal alterations in 1p36 were investigated in 196 neuroblastoma tumors using fluorescence in situ hybridization. Additionally, by using the same technique, it was determined whether MYCN was amplified in 149 of these. The most frequent finding was a deletion in 1p36, leading to monosomy of this region (29 cases, 15%). Furthermore, we found tumors with at least two intact copies of chromosome 1 and additional 1p36-deleted copies. Altogether, 21 tumors (11%) displayed this imbalance of 1p36. Similar to the cases with deletion, imbalances were predominantly found in stage 4 tumors (81%), and they were significantly associated with an increased patient age (P = 0.01). Nearly all 1p-deleted tumors showed amplification of MYCN (24/27 analyzed samples, 89%), whereas only 8 of 21 (38%) with imbalance did. Eight cases with imbalance were investigated for loss of heterozygosity (LOH) using microsatellite markers in 1p35-36. Only 4 displayed 1p36 LOH, whereas the remaining 4 were heterozygous. Both patients with deletion of 1p and with imbalance had a poor outcome [3-year rate of event-free-survival (EFS): 33 +/- 15% and 41 +/- 15%], which was significantly worse compared to the outcome of patients without 1p alterations (3-year EFS: 70 +/- 5%; P = 0.01 and P = 0.0059). We conclude that besides monosomic short arm deletions, imbalance of 1p36 is a strong marker of a poor prognosis in neuroblastoma and not necessarily associated with MYCN amplification and LOH.     

Interstitial deletion of the long arm of chromosome 11 determined by fluorescencein situ hybridization

Summary An interstitial deletion, del(11)(q14q22), found in a female infant was examined by fluorescencein situ hybridization with cosmid DNA markers mapped on the long arm of chromosome 11. Three cosmids mapped on 11q14.1-11q22.1 region were not hybridized to the del(11) chromosome, while all the other DNA markers mapped on 11cen-11q14.1 and 11q23.1-11 qter region gave hybridization signals on the del(11) chromosome. Cytogenetic analysis after R-banding confirmed an apparent deletion of 11q14-q22, but containing a small R-negative band, a part of 11q22.3 and/or 11q14.1, in the middle part of del(11) chromosome. The karyotype thus was determined to be 46, XX, del(11)(q14.1q22.3).     

Analysis of deletions of the long arm of chromosome 11 in hematologic malignancies with fluorescence in situ hybridization

We studied samples containing deletions of the long arm of chromosome 11 (11 q) from patients with hematologic malignancies by using cytogenetic and fluorescence in situ hybridization (FISH) techniques. Cytogenetic analysis of 28 patients and of a cell line showed that all deletions included band 11q23. FISH analysis demonstrated that the proximal part of 11q23, including NCAM, was deleted in 13 of 15 patients and the cell line. Recurring chromosomal losses in human tumors have been regarded as evidence that the affected regions contain tumor-suppressor genes. These results suggest that the putative tumor-suppressor gene is proximal to the MLL gene which is also located in 11q23. © 1993 Wiley-Liss, Inc.     

Fluorescence in situ hybridization evaluation of chromosome deletion patterns in prostate cancer.

Various nonrandom chromosomal aberrations have been identified in prostate carcinoma. These aberrations include deletions of several chromosome regions, particularly the chromosome 8 short arm. Large-scale numerical aberrations, reflected in aberrant DNA ploidy, are also found in a minority of cases. However, it is unclear whether prostate carcinomas contain aberrations of certain chromosome regions that are deleted frequently in other common types of cancer. In this study, we performed dual-color fluorescence in situ hybridization on intact nuclei from touch preparations of 16 prostate cancers. Chromosome copy number was determined using pericentromeric probes, whereas potential chromosome arm deletions were evaluated using yeast artificial chromosome (YAC) and P1 probes. Two YAC probes targeted chromosome 8 short arm regions known to be deleted frequently in prostate cancer. Other YACs and P1s were for chromosome regions, including 1p22, 3p14, 6q21, 9p21, and 22q12, that are deletion targets in a variety of cancers although not extensively studied in prostate cancer. Hybridization efficiencies and signal intensities were excellent for both repeat sequence (alpha-satellite) and single, copy (YAC and P1) fluorescence in situ hybridization probes. Of 16 prostate cancers, 11 had clonal aberrations of 1 or more of the 13 chromosome regions evaluated, and 10 cases (62.5%) had 8p deletions, including 4 cases with 8p deletion in virtually all cells and aneuploidy in only a subset of those deleted cells. Deletions at 3p14, 6q21, and 22q12 were identified in 2, 1, and 1 case, respectively, and each of those cases had a similarly sized cell population with 8p deletion. These studies confirm 8p deletion in the majority of prostate carcinomas. 8p deletions appear to be early events in prostate tumorigenesis, often antedating aneuploidy. Fluorescence in situ hybridization strategies incorporating pericentromeric and single-copy regional chromosome probes offer a powerful and efficient means for determining frequency and progression of oncogenetic events in prostate cancer.     

A patient with an interstitial deletion of the short arm of chromosome 6

The clinical history and subsequent progress of a child with an interstitial deletion in the short arm of chromosome 6 is described. This abnormality coupled with a reduced Hageman factor (Factor XII) led to an earlier publication which suggested that this gene was localized to the breakpoint region involved. A review of similar phenotypes from the literature is presented.     

Hemizygous deletions of chromosome band 16q24 in Wilms tumor: detection by fluorescence in situ hybridization

Loss of heterozygosity (LOH) for markers on chromosome arm 16q in Wilms tumor has been linked to an increased risk of treatment failure. We therefore postulated that fluorescence in situ hybridization (FISH) with probes from this region might enhance current strategies for identifying high-risk patients at diagnosis. In a blinded comparative pilot study of 19 Wilms tumor samples from 18 patients with favorable histology, FISH and DNA polymorphism analysis yielded concordant results in 14 cases, either retention (n = 6) or loss (n = 8) of chromosome arm 16q markers. Discordant findings in 4 of the 5 remaining cases resulted from detection of LOH, but no loss by FISH. Two of these cases, directly comparable at marker D16S422, appeared to have tumor-specific uniparental disomy, in that 2 copies of D16S422 and the 16 centromere were evident, despite LOH. In 2 other cases, the discrepancies could be explained by LOH confined to loci distal to the D16S422 locus. In the fifth case, FISH detected 2 distinct populations of tumor cells, one characterized by normal diploidy and the other by monosomy 16, whereas DNA polymorphism analysis failed to indicate LOH altogether. Thus, FISH confirmed the presence of allelic loss (hence, the possible location of biologically important tumor suppressor genes) on the distal long arm of chromosome 16 in cases of favorable-histology Wilms tumor, with the advantages of technical simplicity, successful analysis of samples that were otherwise uninformative by analysis of DNA polymorphisms, and the addition of internal controls for chromosomal aneusomy. We suggest that combined analysis of the chromosome 16q region in Wilms tumor by FISH and DNA polymorphism analysis would improve evaluations to identify high-risk patients who might benefit from alternative therapy.     

Mapping of a gene causing familial Mediterranean fever to the short arm of chromosome 16.

BACKGROUND. Familial Mediterranean fever is an autosomal-recessive disease characterized by acute attacks of fever with sterile peritonitis, pleurisy, or synovitis. The biochemical basis of the disease is unknown, but determining the chromosomal location of the gene for the disorder should be a first step toward defining the biochemical events. METHODS AND RESULTS. As part of a systematic genome-wide search, we sought evidence of linkage between familial Mediterranean fever and chromosome 16 DNA markers in 27 affected non-Ashkenazi Jewish families from Israel. Two loci from the subtelomeric region of the short arm of chromosome 16 (16p) had lod scores sufficient to establish linkage (a score greater than or equal to 3). One DNA marker (D16S84) gave a maximal lod score of 9.17 (odds of 10(9.17) to 1 in favor of linkage) at a recombination frequency (theta) of 0.04. A probe associated with the hemoglobin alpha complex (5'HVR) gave a maximal lod score of 14.47 at a theta of 0.06. Multipoint linkage analysis indicated that the following was the most likely gene order: the centromere, the gene for familial Mediterranean fever, D16S84, hemoglobin alpha, and the telomere. The maximal multipoint lod score was 19.86. There was a striking degree of homozygosity at chromosome 16p loci in the affected offspring of eight consanguineous couples. CONCLUSIONS. The gene that causes familial Mediterranean fever in non-Ashkenazi Jews maps to the short arm of chromosome 16.     

Fluorescence in situ hybridization analysis of chromosome 12 anomalies in semen cells from patients with carcinoma in situ of the testis

Carcinoma in situ (CIS) of the testis is the precursor of seminomas and non-seminomatous germ cell tumours of the adult testis. A marked cytogenetic anomaly, the isochromosome of the short arm of chromosome 12 [i(12p)], has been demonstrated in over 80 per cent of all histological varieties of testicular germ cell tumours (TGCTs). In the remaining group of i(12p)-negative TGCTs, an overrepresentation of chromosome 12p sequences has been found. The i(12p) chromosome and overrepresentation of 12p sequences in CIS cells have also been reported. In order to establish whether numerical and/or structural aberrations of chromosome 12 can be found in CIS cells exfoliated into seminal fluid, semen specimens from ten patients with CIS lesions were investigated using bicolour double fluorescence in situ hybridization (FISH). The two DNA probes used, p alpha 12H8 and YAC 5, specifically detect the centromeric region of chromosome 12 and a subregion, p11.2-p12.1, on the short arm of chromosome 12, respectively. Ejaculates of ten azoospermic or oligozoospermic infertile males, presumably CIS-free, were used as negative controls. Nuclei exhibiting three or more chromosome 12 signals were found to be present in a significantly larger number in the patient samples than in the control samples. Nuclei with five or more chromosome 12 signals were observed in eight out of the ten patients. Morphologically similar arrangements to i(12p) were observed in some of the ejaculates. These results demonstrate the potential of FISH in the early detection of CIS and TGCTs in males at high risk.     

Preferential involvement of the short arm in chromosome 8‐derived supernumerary markers and ring as identified by chromosome arm painting

We report on the use of fluorescence in situ hybridization (FISH) with specific chromosome 8 arm painting to characterize further small supernumerary chromosome 8‐derived markers/rings (SMC/SRC) identified in three patients. Two patients (patients 1 and 2) who carried the marker (SMC) were evaluated because of mental retardation and minor facial anomalies. The patient (patient 3) who carried the ring (SRC) had ventriculomegaly. Parental blood chromosomes of patients 2 and 3 were normal and unavailable on patient 1. The identification of the SMC/SRC was first characterized by FISH specific alpha‐repeat centromeric probes, second by FISH whole chromosome painting (WCP), and finally by FISH chromosome arm painting (CAP). The latter showed involvement of only the short arm of chromosome 8 in all three SMC/SRC cases, suggesting a U‐type exchange mechanism. Am. J. Med. Genet. 90:276–282, 2000. © 2000 Wiley‐Liss, Inc.     

Fluorescence in situ hybridization for the study of cell lineage involvement in myelodysplastic syndromes with chromosome 5 anomalies

Fluorescence in situ hybridization (FISH) with a locus-specific dual DNA probe (LSI EGR-1SO/D5S23SG) for chromosome 5 was used in combination with morphology to study bone marrow cell lineage involvement of the abnormal chromosomal clone in 13 patients with deletion 5q [del(5q)], either as a sole aberration or as part of a complex karyotype, and in six cases with monosomy 5 by metaphase cytogenetics, all with complex karyotypes including 2-6 marker chromosomes. In the monosomy 5 group, only one case displayed the expected one orange and one green (1O + 1G) FISH pattern in a majority of the cells. The other five patients instead showed 1O + 2G FISH signals in 17-86% of the bone marrow cells, which is the typical pattern for del(5q). In the del(5q) group, 26-98% of the bone marrow cells exhibited 1O + 2G FISH signals. All patients showed clonal involvement of the myeloid cell lineages, including the megakaryocytes in a few cases, whereas lymphoid cells generally exhibited the normal 2O + 2G FISH pattern. No difference was seen between patients with 5q- syndrome, those with del(5q) and a complex karyotype, and the monosomy 5 group. We were thus unable to confirm the recent suggestion that B-cells are a part of the abnormal clone in MDS with del(5q). Furthermore, true monosomy 5 seems to be rare in MDS.