No increased chromosome breakage in skin fibroblasts from patients with musculoskeletal sarcoma

The frequency of structural chromosome aberrations was studied in cultured skin fibroblasts from 25 untreated patients with musculoskeletal sarcoma and 25 controls. Subepidermal skin biopsies from the sarcoma patients were taken either close to the tumor (10 patients), at a distance from the tumor, i.e., from a contralateral limb (8 patients), or from both sites (7 patients). The aberration frequencies in fibroblasts obtained from different locations did not differ significantly (P greater than 0.05). The two groups of fibroblast lines were therefore pooled, yielding a total of 25 sarcoma patients with the following mean frequencies of aberrant cells, and gap, break, and gap + break events: 5.3, 2.8, 3.2, and 6.1. Among the controls the corresponding rates were 5.1, 2.6, 3.2, and 5.8. The differences were not statistically significant. These results provide no evidence of local differences in chromosome aberration frequencies, or that inherent chromosomal instability is important for the pathogenesis of sarcomas.     

Mutagen sensitivity as an indicator of soft tissue sarcoma risk.

The etiology of soft tissue sarcoma is poorly understood. Exposure to environmental chemicals may play a role, but the data are not clear. We compared a group of soft tissue sarcoma patients with healthy controls to determine whether the mutagen sensitivity assay, a simple chromosome aberration assay using the radiomimetic bleomycin, might be useful to identify patients at risk for soft tissue sarcoma. Patients with a diagnosis of soft tissue sarcoma at Memorial Sloan-Kettering's outpatient clinic signed informed consent and donated 30 ml of blood. Controls were selected from the general population of Connecticut by random digit dialing. Unrepaired DNA damage was assessed for 100 metaphase spreads for each individual, with the number of breaks in chromatids being counted as breaks per cell (b/c). The 20 cases with soft tissue sarcoma had 1.03 mean b/c and the controls had 0.88 b/c (P = 0.16). Patients with soft tissue sarcoma were 5.7 times more likely to be mutagen sensitive than controls (P = 0.01), as determined after dividing subjects into sensitive or not sensitive groups based on the median b/c among controls. As mutagen sensitivity has been shown to be associated with a number of cancers and appears to reflect genetic susceptibility, this assay may be an appropriate biomarker for radiation sensitivity or it may be a marker of susceptibility to soft tissue sarcoma. Larger studies should be undertaken to assess these possibilities.     

Chromosome fragility in Alzheimer's disease

We present cytogenetic findings for 12 patients with Alzheimer's Disease (AD) mean age 75.8 ± 6.01 years and 35 normal age and sex matched controls (mean age 74.8 ± 4.04 years). The study, undertaken due to reports of increased fragments and chromosome breakage in individuals with AD, was performed blind on coded peripheral blood specimens and the allocation of AD or control was not known to the cytogenetic staff until the end of the study. Both the AD group and the controls have been very carefully selected and both underwent the same clinical assessment and screening procedures which included CT scanning.
Chromosomes were analysed after 72 h cultures, using deprived medium TC199 which is known to enhance the appearance of fragile sites. A minimum of 50 cells was examined in each case and any rearrangement found was classified as ctg, csg, ctb, csb and the chromosome in which it occurred was recorded. Analysis of results showed that there was no statistically significant difference between the AD group and the controls for either the total occurrence of breaks, the type of aberration or the chromosome(s) involved. In both groups the commonest break was in 3p.     

Chromosome instability in lymphocytes from patients with celiac disease

Cytogenetic studies were performed in celiac disease (CD) patients to determine if the presence of chromosome instability is related to the predisposition to cancer. Chromosome aberrations (CA) and sister chromatid exchange (SCE) frequencies in peripheral blood lymphocyte cultures from untreated CD patients and healthy controls were analyzed. Patients showed aberrations in 23% of cells, while only 3% were detected in the control group (p < 0.0001). The mean frequencies of gaps, breaks and total CA were found to be higher in CD patients compared to controls (p < 0.0001). Breakpoint distribution was nonrandom among chromosomes from celiac patients (p = 0.01), but not among controls (p = 0.04). The frequency of SCE/cell showed a mean value of 6.9 ± 0.6 in CD patients and 7.3 ± 0.2 in controls. No statistical differences were found. Breakpoints involved in CD patients presented a strong coincidence with the location of fragile sites (78.6%) and sites of cancer chromosome rearrangements (57.1%), most of them (75%) associated with malignant non-Hodgkin lymphomas. These results suggest that CD is a condition with increased chromosome instability characterized by a high level of CA and normal SCE frequencies, probably related to the increased incidence of cancer.     

Fluorescence in situ hybridization analysis of chromosome 12p in paraffin-embedded tissue is useful for establishing germ cell origin of metastatic tumors.

The over-representation of chromosome 12p sequences is crucial for the development of invasive testicular germ cell tumors. Testicular cancer patients may have metastatic tumors of diverse histologic types, including adenocarcinoma, undifferentiated carcinoma, sarcoma, or other malignancies that lack features of germ cell tumors. We sought to investigate the possible germ cell origin of such tumors using interphase fluorescence in situ hybridization. In all, 10 metastatic malignant somatic-type tumors from patients with histories of testicular cancer, as well as one malignant somatic-type tumor from a patient with primary mediastinal germ cell tumor were studied and included: adenocarcinoma (five cases), poorly differentiated carcinoma (one), sarcoma (four), and neuroendocrine carcinoma (one). The tumors were analyzed using fluorescence in situ hybridization using 12p spectrum green and 12 centromeric spectrum orange probes in paraffin sections. The patients ranged in age from 27 to 55 years (mean, 43). Colon and lung cancers from patients without germ cell tumors were used as controls. Adequate signals were observed in all tumors. Gain of chromosome 12p was seen in six tumors. None of the control tumors showed 12p amplification. Fluorescence in situ hybridization for 12p amplification in routinely processed surgical specimens is a useful adjuvant diagnostic tool in confirming the germ cell origin of metastatic tumors having the histologic appearance of somatic-type neoplasms.     

A possible fragile-site at Yq12: case report and possible relevance to de novo structural rearrangements of the Y-chromosome

In a routine chromosome study it was noted that cells from a patient had different lengths of Y-chromosome: 20% of the cells had a Y-chromosome with about 40% of the normal length of heterochromatin, whereas the majority of cells had an average size Yq12 region. A break within Yq12 was found in 3-4% of the cells with the long Yq12 chromosome, indicating the presence of a possible new fragile site. The frequency of cells with the 2 different sizes of Y-chromosomes was unaffected by culture conditions, as was the frequency of cells with a break or gap. Possible consequences of a fragile-site within Yq12 are discussed.     

Use of interphase fluorescence in situ hybridization in prostate needle biopsy specimens with isolated high-grade prostatic intraepithelial neoplasia as a predictor of prostate adenocarcinoma on follow-up biopsy

Isolated high-grade prostatic intraepithelial neoplasia (HGPIN) on needle biopsy confers an increased risk of prostate carcinoma (CaP) on follow-up biopsy. The aim of this study is to determine whether paraffin-section fluorescence in situ hybridization (FISH) of specific chromosome/oncogene copy number abnormalities (CNAs) in biopsy specimens with isolated HGPIN increases the predictive value for CaP on repeat biopsy. Cases were divided into 3 groups: controls (n=8) and sextant biopsy specimens with isolated HGPIN without CaP (group A; n=11) and with CaP (group B; n=14) on follow-up biopsy. Dual-color FISH assessing c-myc, HER-2/neu, chromosome region 7q31 (D7S486), and corresponding chromosome centromeres was performed. An amplification ratio (AR) for each marker centromere was derived for each biopsy specimen, with AR ranges designated as no/low, low-intermediate, and high. Also calculated for each marker were the percentage of cells with marker amplification, hyperdiploidy, and monosomy. A composite score for each biopsy specimen was calculated based on these parameters, with a possible range of 0 to 15. The specific chromosomal oncogene CNAs were as follows: for chromosome 7/7q31, 2 of 11 (18%) in group A and 6 of 14 (43%) in group B; for chromosome 8/c-myc, 4 of 11 (36%) in group A and 9 of 13 (69%) in group B; and for chromosome 17/HER-2/neu, 10 of 10 (100%) in group A and 13 of 14 (93%) in group B. The mean composite score was 0 for controls, 2.5 for group A, and 4.7 for group B. Composite scores > or =4 for the 3 groups were 0 of 9 (0%) for controls, 1 of 11 (12%) for group A, and 8 of 14 (57%) for group B. These differences were statistically significant (P=0.015). One group A patient with a high composite score (6) had atypical small glands on follow-up biopsy at <1 year. Chromosome/oncogene CNAs are uncommon in control patients, occurring with increasing frequency and magnitude in patients with isolated HGPIN without and with follow-up CaP. Chromosome/oncogene CNAs in HGPIN are mostly of the low to intermediate level and display intercellular heterogeneity. HER-2/neu amplification is common in HGPIN with and without follow-up CaP. Chromosome 7 and 8 aneusomy and 7q31 and c-myc amplification are greater in HGPIN with follow-up CaP. Patients with isolated HGPIN and high composite score without follow-up CaP are uncommon; these patients may have a small, unsampled CaP. Although patients with HGPIN without CaP are more likely to have a low composite score, a subset of patients with follow-up CaP have low composite score, suggesting (1) mutational pathways independent of chromosomes 7, 8, and 17 and HER-2/neu, c-myc, and chromosome region 7q31 CNAs; (2) CaP derived from an independent, unsampled focus of HGPIN; or (3) CaP not derived from HGPIN.     

Chromosome aneuploidy in Alzheimer''s disease

We present cytogenetic findings in 9 patients with Alzheimer's disease (AD) and 35 normal age matched controls. The study was undertaken due to reports of increased aneuploidy in individuals with AD. Coded peripheral blood chromosome preparations were evaluated for aneuploidy; there were 4.5% hypomodal cells in AD patients compared with 5.5% in controls. There were no hypermodal cells in the AD group and only 0.7% among the controls. Statistical analyses of the results did not show any differences between AD patients and controls in these data. There was a statistically significant increase in the loss of C group chromosomes in the old controls which we attribute to ageing.     

间期双色双融合荧光原位杂交技术检测成人B细胞急性淋巴细胞白血病费城染色体

目的 探讨成人B细胞急性淋巴细胞白血病(B-lineage acute lymphoblastic leukemia.BALL)中费城染色体(Philadelphia chromosome,Ph染色体)的发生率.方法 应用针对BCR-ABL的双色双融合探针和间期荧光原位杂交(dual-color dual-fusion interphase fluorescence in situ hybridization,DDFISH)技术前瞻性检测112例初诊成人B-ALL患者染色体标本中Ph染色体.并与常规细胞遗传学(conventional cytogenetics,CC)结果 进行比较.结果 成功进行CC检测的89例患者中有16例(17.98%)检出Ph染色体,而DD-FISH检测112例发现35例(31.25%)Ph染色体,阳性细胞率为18.50%～99.00%(平均66.23%).35例DD-FISH检测出Ph+的成人B-ALL患者中,10例未能成功进行CC检测,5例核型正常,16例检出Ph染色体,20例核型异常患者中13例为复杂异常.结论 Ph染色体在成人B-ALL中以DD-FISH检测发生率为31.25%;用BCR-ABL探针进行DD-FISH为检测Ph染色体提供了有效的方法 .是细胞遗传学检查的重要补充;对成人B-ALL患者应常规进行DD-FISH检测.     

Sister chromatid exchange in malignant lymphomas

Sister chromatid exchange (SCE) was evaluated in peripheral lymphocytes from 20 untreated patients with malignant lymphomas: 6 with Hodgkin's disease (HD), 14 with non-Hodgkin lymphoma (NHL), and 5 with lymphadenitis. The mean SCE frequency (+/- SE) was: 11.2 +/- 0.6, 11.0 +/- 0.6, and 7.2 +/- 0.3 for HD, NHL, and lymphadenitis patients, respectively, and 8.7 +/- 0.2 for the control group. No differences in SCE score were observed in HD and NHL. These results allowed us to consider both groups (HD and NHL) as a single neoplastic population (mean +/- SE, 11.0 +/- 0.4). No significant differences were found between the lymphadenitis and control groups. On the other hand, significantly higher SCE scores were seen in neoplastic populations than in the control and lymphadenitis groups (p less than 0.001 and p less than 0.01, respectively). When SCE was compared by chromosome number and group between neoplastic patients and controls, a higher SCE frequency was observed in chromosomes #1, #2, #3, and B, C + X, E, F chromosome groups than in controls. SCE levels were significantly higher in lymphoma patients in all chromosome numbers and groups mentioned than in patients with lymphadenitis. It is suggested that the high SCE rate in the malignant lymphoma population is possibly related to an increased chromosomal instability.     

Functional disomies of the X chromosome influence the cell selection and hence the X inactivation pattern in females with balanced X-autosome translocations: a review of 122 cases.

We reviewed 122 cases of balanced X-autosome translocations in females, with respect to the X inactivation pattern, the position of the X break point and the resulting phenotype. In 77% of the patients the translocated X chromosome was early replicating in all cells analysed. The break points in these cases were distributed all along the X chromosome. Most of these patients were either phenotypically normal or had gonadal dysgenesis, some had single gene disorders, and less than 9% had multiple congenital anomalies and/or mental retardation. In the remaining 23% of the cases the translocated X chromosome was late replicating in a proportion of cells. In these cells only one of the translocation products was reported to replicate late, while the remaining portion of the X chromosome showed the same replication pattern as the homologous part of the active, structurally normal X chromosome. The analysis of DNA methylation in one of these cases confirmed noninactivation of the translocated segment. Consequently, these cells were functionally disomic for a part of the X chromosome. The presence of disomic cells was highly prevalent in translocations with break points at Xp22 and Xq28, even though spreading of X inactivation onto the adjacent autosomal segment was noted in most of these cases. This suggests that selection against cells with a late replicating translocated X is driven predominantly by a functional disomy X, and that the efficiency of this process depends primarily on the position of the X break point, and hence the size of the noninactivated region.(ABSTRACT TRUNCATED AT 250 WORDS)     

Functional disomies of the X chromosome influence the cell selection and hence the X inactivation pattern in females with balanced X-autosome translocations: A review of 122 cases

We reviewed 122 cases of balanced X-autosome translocations in females, with respect to the X inactivation pattern, the position of the X break point and the resulting phenotype. In 77% of the patients the translocated X chromosome was early replicating in all cells analysed. The break points in these cases were distributed all along the X chromosome. Most of these patients were either phenotypically normal or had gonadal dysgenesis, some had single gene disorders, and less than 9% had multiple congenital anomalies and/or mental retardation. In the remaining 23% of the cases the translocated X chromosome was late replicating in a proportion of cells. In these cells only one of the translocation products was reported to replicate late, while the remaining portion of the X chromosome showed the same replication pattern as the homologous part of the active, structurally normal X chromosome. The analysis of DNA methylation in one of these cases confirmed noninactivation of the translocated segment. Consequently, these cells were functionally disomic for a part of the X chromosome. The presence of disomic cells was highly prevalent in translocations with break points at Xp22 and Xq28, even though spreading of X inactivation onto the adjacent autosomal segment was noted in most of these cases. This suggests that selection against cells with a late replicating translocated X is driven predominantly by a functional disomy X, and that the efficiency of this process depends primarily on the position of the X break point, and hence the size of the noninactivated region. Since the persistence of cells with a late replicating translocated X was usually associated with mental retardation and other abnormalities, it is concluded that the outcome of the selection process against the functional disomy X is the major determinant of the clinical status in most patients with balanced X-autosome translocations.     

The expression frequency of common fragile sites and genetic susceptibility to lung cancers

The chromosomal aberration rate (including gap and break) and expression frequency of common fragile sites were examined in peripheral blood lymphocytes cultured with TC199 medium from 96 patients with lung cancers, 40 of their first-degree relatives, and 45 normal control subjects. Both the chromosomal aberration rates and expression frequencies of common fragile sites observed in patients and their relatives were significantly higher than those in normal control subjects. About 60% of chromosomal aberrations were derived from the expression of common fragile sites either in the patients and their relatives or in the controls. The expression of fra(3)(p14) was most frequently observed, and the mean frequencies of its expression in patients and their relatives were significantly higher than in control subjects. It is suggested that common fragile sites might be unstable factors of the human genome, and their expression might be affected by some genetic factors, and they might play an important role in the genetic susceptibility to lung cancers. The significantly high expression of fra(3)(p14) in patients and their relatives may be related to the generation of the breakpoint at band 3p14 found in lung cancers.     

Ewing's sarcoma: diagnostic,prognostic, and therapeutic implications of molecular abnormalities

The identification of the non-random chromosome rearrangements between the EWS gene on chromosome 22q12 and members of the ETS gene family in Ewing's sarcoma, peripheral primitive neuroectodermal tumour, Askin tumour, and neuroepithelioma has been a key advance in understanding their common histogenesis and defining the Ewing's sarcoma family of tumours (ESFT). In addition to improvements in diagnosis and potentially the stratification of patients for risk, biological investigations of these gene fusions may define targets for much needed therapeutic strategies to eliminate minimal residual disease or metastatic disease. Insight into their relation with other oncogenic events in ESFT will advance risk group analysis and ultimately may improve clinical management and survival for patients with this disease.     

Detection of aneuploidy for chromosomes 4, 6, 7, 8, 9, 10, 11, 12, 13, 17, 18, 21, X and Y by fluorescence in-situ hybridization in spermatozoa from nine patients with oligoasthenoteratozoospermia undergoing intracytoplasmic sperm injection.

Recent evidence suggests that infertile males donating semen for intracytoplasmic sperm injection (ICSI) may be at an increased risk of transmitting numerical (predominantly sex chromosome) abnormalities to their offspring. The present study was designed to determine aneuploidy in spermatozoa from oligoasthenoteratozoospermic (OAT) patients undergoing ICSI. Aneuploidy frequencies of 12 autosomes and the sex chromosomes were determined by fluorescence in-situ hybridization (FISH) on spermatozoa from fresh ejaculate of nine severe OAT patients and four proven fertile donors. FISH, using directly labelled (fluorochrome-dUTP) satellite or contig DNA probes specific for chromosomes 4, 6, 7, 8, 9, 10, 11, 12, 13, 17, 18, 21, X, and Y, was performed on decondensed spermatozoa. Per chromosome disomy frequencies for autosomes and sex chomosomes in OAT males were 0-5. 4%. In contrast, the disomy frequencies in controls were 0.05-0.2%. The frequency of diploid spermatozoa in OAT patients was 0.4-9.6%; controls showed a mean of 0.04%. Using recently developed formulae, the total aneuploidy in our OAT patient population was estimated to be 33-74%. In contrast, estimates of mean total aneuploidy in the spermatozoa of controls ranged from 4.1 to 7.7%, depending upon method of calculation. Six series of ICSI were performed on five of the OAT patients. Four resulted in no establishment of pregnancy; the others failed to establish ongoing pregnancies. Our cytogenetic data show significantly elevated frequencies of diploidy, autosomal disomy and nullisomy, sex chromosome aneuploidy, and total aneuploidy in OAT patients, which may contribute to the patients' infertility.     

Analysis of premature centromere division (PCD) of the chromosome 18 in peripheral blood lymphocytes in Alzheimer disease patients

Premature centromere division (PCD) of the chromosome 18 was analyzed by using fluorescent in situ hybridization (FISH) on interphase peripheral blood lymphocytes isolated from six sporadic Alzheimer disease (AD) patients and six healthy elderly controls. Results of FISH analysis revealed that chromosome 18 expressed PCD in 5.18% interphase nuclei of AD patients, and in 2.59% interphase nuclei of age-matched controls (p<0.05). Our study also showed that hypoploidy and hyperploidy frequency for chromosome 18 exhibited a statistically significant increase in the AD group compared to the control one. The increase in spontaneous aneuploidy of chromosome 18 in AD patients which is correlated with PCD shows that deregulation of the time of centromere separation can be considered as a manifestation of chromosome instability leading to aneuploidy.     

Increased skewing of X chromosome inactivation in Rett syndrome patients and their mothers

Rett syndrome is a largely sporadic, X-linked neurological disorder with a characteristic phenotype, but which exhibits substantial phenotypic variability. This variability has been partly attributed to an effect of X chromosome inactivation (XCI). There have been conflicting reports regarding incidence of skewed X inactivation in Rett syndrome. In rare familial cases of Rett syndrome, favourably skewed X inactivation has been found in phenotypically normal carrier mothers. We have investigated the X inactivation pattern in DNA from blood and buccal cells of sporadic Rett patients (n=96) and their mothers (n=84). The mean degree of skewing in blood was higher in patients (70.7%) than controls (64.9%). Unexpectedly, the mothers of these patients also had a higher mean degree of skewing in blood (70.8%) than controls. In accordance with these findings, the frequency of skewed (XCI > or =80%) X inactivation in blood was also higher in both patients (25%) and mothers (30%) than in controls (11%). To test whether the Rett patients with skewed X inactivation were daughters of skewed mothers, 49 mother-daughter pairs were analysed. Of 14 patients with skewed X inactivation, only three had a mother with skewed X inactivation. Among patients, mildly affected cases were shown to be more skewed than more severely affected cases, and there was a trend towards preferential inactivation of the paternally inherited X chromosome in skewed cases. These findings, particularly the greater degree of X inactivation skewing in Rett syndrome patients, are of potential significance in the analysis of genotype-phenotype correlations in Rett syndrome.     

Sister chromatid exchanges in leukemic patients

Sister chromatid exchange (SCE) was studied in PHA-stimulated peripheral blood lymphocytes from 36 newly diagnosed and untreated leukemic patients: 16 with acute lymphoblastic leukemia (ALL), 10 with acute nonlymphocytic leukemia (ANLL), and 10 with chronic myelocytic leukemia (CML). The metaphases analyzed show no chromosomal abnormalities. The mean SCE frequency (mean +/- SE) for each group of patients was: 6.8 +/- 0.4, 6.6 +/- 0.3, and 7.0 +/- 0.6 per mitosis, respectively, which was significantly lower than the mean SCE score for 30 controls (8.7 +/- 0.2). No differences in SCE score among ALL, ANLL, and CML and a similar SCE frequency by chromosome number and group allowed consolidation of all the cases into a single group of 36 leukemic patients (6.8 +/- 0.3). When the frequency of SCE was compared by chromosome number and group between the leukemic patients with the control group, a significant decrease in SCE frequency was observed due to a low SCE score in almost all the complements, except chromosome #1. It is suggested that the low SCE rate is related to the leukemic process itself.     

Acquired immune dysfunction in homosexual men: immunologic profiles

Homosexual men with Kaposi sarcoma, lymphadenopathy syndrome, opportunistic infection, and nonhomosexual traditional Kaposi sarcoma were evaluated for B cell, T cell, and complement immunity and compared to normal controls and homosexual controls. No significant immunologic abnormalities were found in the traditional Kaposi group. All homosexual groups, including the homosexual controls, had a significant decrease in the helper/suppressor cell ratio. Functional abnormalities of T-cell immunity were observed in the homosexual Kaposi sarcoma, lymphadenopathy syndrome, and opportunistic infection groups. Significant elevations of IgG, IgM, IgA, and IgE were found in the lymphadenopathy group, while only IgG and IgA were elevated in the Kaposi sarcoma group. C3, C4, and immune complexes were normal, while total hemolytic complement activity was increased in the Kaposi sarcoma and lymphadenopathy syndrome groups.