Deletion of chromosome region 13q14 is transmissible and does not always predispose to retinoblastoma

Summary During routine screening of retinoblastoma patients for esterase D activity in red blood cell lysates a patient was identified with only 50% of normal enzyme activity. Chromosome analysis showed that this patient had a small deletion within chromosome region 13q14. Parental studies showed that, whereas the father had normal enzyme levels, the mother had esterase D levels which were also 50% of normal and a similar small 13q14 deletion. Ophthalmological examination failed to demonstrate any retinal abnormality in either parent. Thus wer present the first case not only of the direct transmission of a 13q14 deletion within a family but also of an individual in whom the deletion has not predisposed to tumour formation.     

Familial retinoblastoma (mother and son) with 13q14 deletion

Summary We present here the first familial cases (a mother and son) of dominantly inherited retinoblastoma with a 13q14 deletion [46,XY or XX,del(13)(q14.1q21.2)]. Their esterase D activities in red blood cells were as low as 50% of the normal control and the haplotype of esterase D was a type 1-0 in the mother and a type 2-0 in the son. They had peculiar facies characterized by a high forehead, low and broad nasal root, a short and bulbous nose, a long philtrum, and open mouth with a thin upper lip, and prominent earlobes. Chromosome and esterase D analysis should be performed in patients with retinoblastoma even if retinoblastoma seems to be transmitted through an autosomal dominant inheritance. This family indicates that one of the causes of dominantly inherited retinoblastoma is a chromosome deletion of part of the 13q14 band whether it is detectable by chromosome analysis or not.     

Deletions of the esterase D locus from a survey of 200 retinoblastoma patients

Summary Esterase D levels from 200 retinoblastoma patients have been measured in an attempt to identify individuals carrying deletions of chromosome region 13q14. In this series 75% had bilateral tumours and 23% were familial. Of nine patients identified as having low esterase D levels, five had not previously been diagnosed as deletion carriers. These observations demonstrate the benefit of screening retinoblastoma populations for esterase D deficiency.     

A null allele of esterase D is a marker for genetic events in retinoblastoma formation

Summary The development of homozygosity or hemizygosity in the 13q14 region by deletion, mitotic recombination, or chromosomal loss has been interpreted as a primary event in retinoblastoma. This finding is consistent with the hypothesis that inactivation of both alleles of a gene located at 13q14.11 is required for tumorigenesis. Observations reported by Benedict and colleagues in one case of bilateral retinoblastoma, LA-RB 69, provided early evidence in favor of this hypothesis. By examining levels of esterase D, an enzyme also mapping to 13q14.11, it was previously inferred that one chromosome 13 in this patient's somatic cells contained a submicroscopic deletion of the Rb and esterase D loci and that this chromosome was retained in her tumor while the normal chromosome 13 was lost. Using a rabbit anti-esterase D antibody and the esterase D cDNA probe, we have found that (1) low but detectable quantities of esterase D protein and enzymatic activity are present in tumor cells from LA-RB 69; (2) fibroblast from this patient contain two copies of the esterase D gene, indicated by heterozygosity at an ApaI polymorphic site within this gene; and (3) tumor cells from the same patient are homozygous at this site, indicating loss and reduplication of the esterase D locus. These results demonstrate that one of the two esterase D alleles in this patient acted as a null or silent allele — that is, was present in the genome with markedly decreased protein expression. This mutant allele acted as a marker for tumor-associated loss of chromosome 13 heterozygosity, in concordance with previous proposals.     

Re-evaluation of the sublocalization of esterase D and its relation to the retinoblastoma locus by in situ hybridization

In situ hybridization of a cDNA probe for the esterase D gene (ESD) was carried out on human chromosomes. The probe hybridized most strongly to 13q14.2 and 13q14.3. This observation raises doubts concerning the most recently published assignment of ESD to 13q14.1. A deletion in an individual with retinoblastoma was reported to separate the closely linked ESD and retinoblastoma (RB1) loci, placing ESD proximal to RB1. Quantitative in situ hybridization studies of this deletion do not confirm this interpretation. Rather, they suggest that ESD is missing from the deleted chromosome 13 and duplicated on the normal homolog. From these findings, we conclude that the deletion in this individual cannot be used to determine the orientation nor the sublocalization of ESD and RB1 within the 13q14 region.     

Interstitial deletion of chromosome 13 involving the region 13q14

Summary A patient with an interstitial deletion 13q14 is described who has decreased erythrocyte esterase D activity and who has not developed a retinoblastoma.     

De novo complex autosomal translocation involving chromosomes 8, 13 and 15 in a girl with a sporadic retinoblastoma

Summary We report a case of a 5-month-old female with sporadic monolateral retinoblastoma (RB) with a constitutional de novo complex autosomal translocation involving chromosomes 8, 13 and 15 resulting in a deletion of chromosome 13q14 confirmed by esterase D assay. The translocation of the terminal portion of chromosome 8 has been observed by in situ hybridization with c-myc and thyroglobulin probes.     

Molecular evidence that the esterase-D gene lies proximal to the retinoblastoma susceptibility locus in chromosome region 13q14

Summary Somatic cell hybrids have been created between transformed mouse 3T3 cells and fibroblasts from a retino-blatoma patient with normal red-cell esterase-D (ESD) levels and a constitutional deletion of chromosome region 13q14-q31. In one subclone, which has retained the deletion chromosome but not the homologous normal copy, we have demonstrated the presence of the human ESD gene sequence. The breakpoint in this patient therefore must have occurred between the ESD gene and the retinoblastoma (Rb) predisposition locus. We have also been able to demonstrate that the ESD gene lies proximally to be the Rb gene in region 13q14. The recently isolated 4.7R cDNA gene sequence was absent from the deletion-containing hybrid, a finding consistent with the hypothesis that this sequence represents the Rb gene itself.     

High rate of detection of 13q14 deletion mosaicism among retinoblastoma patients (using more extensive methods)

Summary Using a cell population with a high proportion of early mitotic cells and by examining more cells derived from peripheral lymphocytes, we found three cases with a 13q14 deletion mosaicism among fifteen retinoblastoma patients; one with a de novo 13/18 balanced translocation, and another with a monosomy 13(q13»q21.2 or 21.3). The three patients with a 13q14 deletion mosaicism had sporadic retinoblastoma (two had bilateral and one unilateral retinoblastoma). The results indicate that 13q14 deletion mosaicism plays a major role in the etiology of this tumor.     

Sister chromatid exchanges and chromosome aberrations in fibroblasts from patients with retinoblastoma

Summary The frequencies of sister chromatid exchanges (SCEs) and chromosome breaks were investigated in five diploid fibroblast strains derived from three patients with deletion 13[del(13)] retinoblastoma, one patient with a hereditary form of retinoblastoma, and one trisomy 13. The fibroblasts with del(13)(q14q22) showed slightly increased SCEs (at a P level of 5–10%), but the others, including del(13)(q12q14), the hereditary form of retinoblastoma, and trisomy 13, did not have increased SCEs as compared to normal controls. No increase in chromosome breaks was found in these fibroblasts. The results suggest that retinoblastoma is not associated with spontaneous increased chromosomal instability.     

Detection of submicroscopic deletions and a DNA polymorphism at the retinoblastoma locus

Summary DNA samples from 60 unrelated patients with retinoblastoma were screened by Southern blot hybridization using two probes that are closely linked to the retinoblastoma locus within human chromosome band 13q14. Seven of 44 patients with bilateral or multifocal unilateral retinoblastoma and one patient with unifocal unilateral retinoblastoma were found to have a heterozygous deletion for the anonymous DNA sequence H3-8. Three of the eight deletions did not include the esterase D locus and were undetectable by conventional cytogenetic analysis. The findings are compatible with the deletions being the cause of retinoblastoma in these cases and provide a basis for DNA diagnosis in nearly 20% of patients with bilateral and multifocal unilateral retinoblastoma. The H3-8 probe also detects a restriction fragment length polymorphism that is a useful genetic marker in some families.     

Retinoblastoma: Host resistance and 13q- chromosomal deletion

Summary Data for 27 cases of retinoblastoma that developed in patients with 13q-were collected from the literature and analyzed. The distribution of unilateral and bilateral cases of retinoblastoma differed significantly from the expectation that the degree of expressivity does not differ between the retinoblastoma gene and deletion of 13q. The excess of unilateral cases among the patients with 13q-, which could not be accounted for by ascertainment bias, was attributed to somewhat lowered carcinogenic potential of deletion of 13q14 as compared with the retinoblastoma gene. It was argued that the retinoblastoma gene is probably not located on 13q, and perhaps 20% or more of the individuals with a deletion of 13q14 would not develop retinoblastoma. The normal allele at the retinoblastoma locus, the haplicon in the segment of 13q14, and the suppressor genes as defined by the host resistance model, may be all concerned, in their function additively and without dominance, with normal differentiation of the embryonic retinal cells.     

A decreasing tendency for cytogenetic abnormality in peripheral lymphocytes of retinoblastoma patients with 13q14 deletion mosaicism

Summary A significant decrease in the proportion of 13q14-deleted cells over a 9-month period was observed in a boy with retinoblastoma and 13q14 deletion mosaicism. To evaluate whether this phenomenon is generally the case, the bloods of three retinoblastoma patients with 13q14 deletion mosaicism reported in 1981 and 1982 were reexamined. A significant decrease in the proportion of abnormal cells was observed in three of four patients including the present case, suggesting that a 13q14 deletion mosaicism might disappear with age in some individuals.     

Translocation (X;13)(p11.21;q12.3) in a girl with incontinentia pigmenti and bilateral retinoblastoma

A de novo t(X;13)(p11.21;q12.3) translocation is described in an 19-month-old girl with incontinentia pigmenti (IP) and bilateral retinoblastoma. Based on previously reported two girls and this patient, each with a structural X chromosome abnormality and IP, it was assumed that the locus for IP is at Xp11.21. Q-banding analysis revealed that the translocated chromosomes were of paternal origin. The derivative X chromosome was late-replicating in 9% of cultured peripheral blood lymphocytes and in 1% of skin fibroblasts. The erythrocyte esterase D activity in the patient was normal. Several possibilities were considered for possible causative relationship between the X/13 translocation and the development of retinoblastoma. One possibility involved functional monosomy of 13q14 in a minority of retinoblasts due to the spreading of inactivation of the translocated X chromosome segment.     

Molecular-genetic analysis of two cases with retinoblastoma: Benefits for disease management

Effective counselling and management of retinoblastoma families using genetic information is presently practised in many parts of the world. We studied histopathological, chromosomal and molecular-genetic data of two retinoblastoma patients from India. The two patients, one with bilateral and the other with unilateral retinoblastoma, underwent complete ophthalmic examination, cytogenetic study, retinoblastoma gene (RB1) mutational analysis andRB1 promoter region methylation screening. In the bilateral retinoblastoma patient deletion of chromosome region 13q14 in peripheral blood lymphocytes and a hemizygous novel 8-bp deletion in exon 4 ofRB1 in tumour sample were observed. In the unilaterally affected patient CGA to TGA transition protein truncation mutations were observed in exons 8 and 14 ofRB1.     

Closely linked loci on the long arm of chromosome 13 flank a specific 2;13 translocation breakpoint in childhood rhabdomyosarcoma

A specific chromosomal translocation, t(2;13)(q35;q14), is present in tumor cells from about one-half of children with alveolar rhabdomyosarcoma, who generally have widely disseminated disease at diagnosis. Using a series of six DNA probes from five loci previously assigned to bands 13q12----q14, we have localized the translocation breakpoint on chromosome 13 by in situ hybridization. Each probe was used to examine metaphase spreads from two or more rhabdomyosarcoma cell lines that have the t(2;13), as well as from control lymphoblastoid cell metaphases. All six probes bound to chromosome 13q12----q14 in the control cell line, but showed no appreciable hybridization to other sites. With rhabdomyosarcoma metaphases, cDNA clones of the retinoblastoma susceptibility gene (RB1) and the esterase D gene (ESD), as well as the arbitrary genomic fragment 7D2 (D13S10), showed specific hybridization to the normal chromosome 13 and the der(2) marker, but not to the der(13). By contrast, the genomic fragments HU10 (D13S6) and 7F12 (D13S1) hybridized specifically to the normal chromosome 13 and the der(13), but not to the der(2). Thus, the breakpoint of this translocation lies distal to D13S6 and D13S1 and proximal to ESD, RB1, and D13S10. Our data indicate that the locus affected by the translocation breakpoint on chromosome 13, which we have termed RMS, is physically distinct from the RB1 locus and is, in fact, proximal to ESD, which others have placed at least 10(6) bp proximal to RB1. The consistent presence of the der(2) marker chromosome, coupled with occasional loss of the der(13), suggests that the RMS gene, or at least a critical component, moves to chromosome 2 in tumors with this translocation.     

Separation of retinoblastoma and esterase D loci in a patient with sporadic retinoblastoma and del(13)(q14.1q22.3)

Summary A chromosome 13 deletion in a patient with sporadic retinoblastoma appears to have separated the loci for retinoblastoma and esterase D. This study indicates that: (1) the retinoblastoma locus is distinct from the esterase D locus; and (2) the linear order of these genes is centromere-esterase D-retinoblastoma.     

RB1 deletion in gonadoblastoma in an XY female

Cytogenetic studies of normal and tumor cells in a patient with gonadal dysgenesis and bilateral gonadoblastoma were performed. The karyotype was 46,XY in peripheral blood lymphocytes and skin fibroblasts. The conserved region of the SRY gene was detected by polymerase chain reaction amplification. Sequencing of this region did not reveal any alterations. A 46,XY chromosome constitution was observed in the right gonadoblastoma, but a partial deletion of chromosome 13 was present in the left tumor. This deletion included band 13q14, where the retinoblastoma gene is mapped. The study of the polymorphism of the variable number of tandem repeats region in intron 17 of the RB1 locus disclosed loss of heterozygosity in both the left tumor, which showed the deletion of chromosome 13, and in the right tumor, where no chromosome alterations of chromosome 13 were detected. In situ hybridization covering 130 kb of RB1 showed that a partial deletion of one of the RB1 alleles had occurred in the right tumor. Since the deletions affected different alleles in each tumor, independent events must have been involved in the development of the tumors. These findings point toward a significant role of RB1 in the development of gonadoblastoma. Received: 17 October 1995 / Accepted: 14 July 1997     

Molecular cytogenetic aberrations in Tunisian patients with multiple myeloma identified by cIg-FISH in fixed bone marrow cells

Cytogenetic studies in multiple myeloma (MM) are hampered by the hypo-proliferative nature of plasma cells. In order to circumvent this problem, we have used a combination of immunolabeling of cytoplasmic Ig light chains (λ or κ) and FISH (cIg-FISH), which allowed a comprehensive detection of the most common and/or recurrent molecular cytogenetic aberrations on fixed bone marrow cells of 70 Tunisian patients. Translocations involving the chromosome 14q32 region were observed in 32 cases (45.7%), including 18 cases with a t(11;14), 8 cases with a t(4;14), and 2 cases with a t(14;16). Deletions of the 13q14 region (D13S319/RB1) were detected in 18.6%, and deletions of the 17p13 region (TP53) in 5.7% of the cases, respectively. Of all patients with a D13S319/RB1 deletion, 61.5% also carried a 14q32 translocation, whereas TP53 deletions were associated with a t(11;14) in 2 cases (50%) and a D13S319 deletion in 1 case (25%). Our results suggest that there is a correlation between the presence of 14q32 translocations and chromosome 13q14 deletions in MM patients and that cIg-FISH is more sensitive as compared to conventional karyotyping in detecting molecular cytogenetic abnormalities in this disease.     

Ultraviolet light and mitomycin C induced sister-chromatid exchanges in fibroblasts from patients with retinoblastoma

Ultraviolet light and mitomycin C (MMC) induced sister-chromatid exchanges (SCEs) were investigated in 6 diploid fibroblast strains derived from 3 patients with deletion 13 and retinoblastoma, one patient with a hereditary form of retinoblastoma, one patient with trisomy 13, and one normal control. Two fibroblast strains with del(13)(q14q22) showed a significant increase in SCEs compared to the control after UV and MMC treatments. In contrast, cell strains with del(13)(q12q14) and trisomy 13 did not show increased SCEs. The frequency of SCEs in fibroblasts from a patient with autosomal dominant retinoblastomas (no deletions) was significantly increased by UV, but not by MMC. The results suggest that cell strains with different deletions of chromosome 13 have different SCE responses to UV and MMC inductions. The cells with del(13)(q14q22) may have a DNA-repair defect.