Deletion (13)(q13q14.3) with retinoblastoma: confirmation and extension of a recognisable pattern of clinical features in retinoblastoma patients with 13q deletion.

A girl with retinoblastoma and a del(13)(q13q14.3) is presented. This case helps to confirm and extend our previous observations regarding a recognisable facial pattern in retinoblastoma patients with 13q deletion involving 13q14 and its adjacent segments.     

Chromosomal anomalies in patients with retinoblastoma

Karyotypes from 50 persons with retinoblastoma confirmed by histopathological examination were studied by conventional staining and Giemsa-banding techniques. Two chromosomal anomalies were found. An interstitial deletion of the long arm of No. 13 chromosome was identified by Giemsa-banding in the karyotypes from a boy with unilateral retinoblastoma. Another boy with unilateral tumor had a karyotype of 47, XXY. These findings provide additional evidence that a deletion of chromosome No. 13, most likely involving band 13q14, is associated with the development of retinoblastoma. In conjunction with other reports, our findings also suggest that retinoblastoma may be found more frequently in children with chromosomal aneuploidy.     

Abnormalities of chromosome #13 in retinoblastomas from individuals with normal constitutional karyotypes

Constitutional chromsome abnormalities have been associated with retinoblastoma, Wilms' tumor, and a familial form of renal cell carcinoma. For each tumor type, the particular chromosome segment involved in the observed rearrangements is different: in retinoblastoma, that segment is band q14 on chromosome #13. We now present evidence that in retinoblastoma, structural abnormalities involving the particular chromosome segment identified in the constitutional cases can also occur in the tumors of individuals with normal constitutional karyotypes. Six cases with retinoblastomas in one or both eyes were analyzed; deletions/rearrangements involving 13q14 were found in the tumor cell karyotypes of five of the six. These observations suggest that changes in a gene or genes at a common site (13q14) play a role in tumorigenesis in all forms of retinoblastoma, sporadic as well as heritable.     

Translocation X;13 in a patient with retinoblastoma.

We describe the clinical and cytogenetic findings in a child with retinoblastoma and a translocation between chromosomes X and 13. The X;13 translocation in this patient does not involve band 13q14, the assigned locus for retinoblastoma.     

A chromosomal breakpoint that separates the esterase D and retinoblastoma predisposition loci in a patient with del(13)(q14q31)

A patient with severe mental retardation and other congenital abnormalities who developed retinoblastoma was shown to have a deletion on the long arm of chromosome #13 with breakpoints in regions q14 and q31. Quantitation of enzyme activity of the esterase-D gene which, together with the retinoblastoma locus, is located in region 13q14 showed levels that were equal to those of normal controls. The 13q14 breakpoint, therefore, appears to have occurred between the two loci, which places the esterase D gene in a more proximal position in this band than the retinoblastoma locus.     

Retinoblastoma and its association with a deletion in chromosome #13: A Survey using high-resolution chromosome techniques

To our knowledge, 20 cases of retinoblastoma associated with a chromosome #13 aberration have been reported. The present study utilized high-resolution prophase banding analysis of 12 additional retinoblastoma patients to determine the occurrence of chromosome aberrations and identify consistently associated clinical abnormalities. Six male and six female patients were studied representing seven cases of bilateral and five cases of unilateral retinoblastoma. One case of unilateral and two cases of bilateral retinoblastoma had detectable cytogenetic abnormalities, all involving an interstitial deletion of 13q14 on the long arm of one chromosome #13. In all five unilateral cases the tumor manifested in the left orbit, and in all seven bilateral cases the left eye was at a more malignantly advanced stage than the right eye. All three cases with a chromosome abnormality had varying degrees of developmental and/or mental retardation, along with at least one other congenital abnormality. In addition to the 12 cases of retinoblastoma, a patient with severe ophthalmologic abnormalities and mild congenital anomalies was studied by the prophase banding technique and found to be partially trisomic for the q13–14 banding regions of chromosome #13. These results further confirm the association of the 13q14 region with gene loci for optic development and indicate that cytogenetic abnormalities may occur even more frequently in retinoblastoma than indicated by the small number of cases reported in the literature.     

Deletion (13)(q14.1q14.3) in two generations: variability of ocular manifestations and definition of the phenotype

A 5-yr-old girl with unilateral retinoblastoma was found to have del(13)(q14.1q14.3). Her 16-month-old sister and 35-year-old mother, with retinal colobomata but without retinoblastoma, have the same deletion. Esterase D studies indicate reduced gene dose at this locus in the 3 females, consistent with a deletion of band 13q14. These patients are of apparently normal intelligence but have a mildly "coarse" facial appearance, a broad nasal bridge, upturned nares, and a long upper lip with thin upper lip vermillion similar to the phenotype suggested by Motegi et al [1983a] for patients with this deletion. Review of the literature documents 2 other patients with deletions of band 13q14 but without retinoblastoma, indicating that retinoblastoma is not a necessary consequence of this deletion. Of the 12 reported patients with deletions limited to band 13q14, seven had normal intelligence and five were macrocephalic. Insufficient clinical information is provided to draw conclusions about phenotype. The family which we describe and those reviewed by Motegi et al suggest that there may be a characteristic appearance in patients with this deletion.     

Chiasma derived genetic maps and recombination fractions: chromosome 13 with reference to the proposed 13q14 retinoblastoma locus.

Provided that there is no chromatid interference, no movement of chiasmata, and no discrepancies between meiotic and mitotic chromosome lengths, then genetic maps and recombination fractions may be directly derived from our meiotic chiasma distribution data. This is illustrated by male chiasma derived genetic lengths and recombination fractions along chromosome 13. The recombination fraction between 13p fluorescent markers and the proposed retinoblastoma locus at 13q14 is estimated at 0.27 to 0.37 and preliminary female chiasma studies suggest a recombination fraction of 0.5 between these two sites. Therefore, it seems unlikely that 13p fluorescent markers may be of any practical help in identifying retinoblastoma gene carriers. This is also borne out by the discordant segregation which has been found in six out of seven retinoblastoma families, which gives a calculated recombination fraction of 0.39 (SE 0.15), not significantly different from 0.5.     

Cytogenetic and molecular investigation of a balanced Xq13q translocation in a patient with retinoblastoma

We report on a 4-year-old girl with retinoblastoma and de novo balanced translocation [46,X,t (X;13) (q23;q13)]. Unilateral retinoblastoma was discovered at age 9 months along with developmental delay and several manifestations of Turner syndrome. Chromosome studies showed an X/13 translocation and an X inactivation pattern showing the translocated X chromosome active in all 50 cells examined. Standard Southern blot analysis and pulsed field gel electrophoresis using a 3.8 kb EcoR1 fragment of the cDNA probe to the 3′ end of the RB1 locus demonstrated a normal genomic pattern. The results of the cytogenetic and molecular analysis suggests that the RB1 locus has not been disrupted by the chromosome rearrangement. This case is the fifth report of an X/13 translocation associated with a retinoblastoma.     

Overexpression and hyperphosphorylation of retinoblastoma protein in the progression of malignant melanoma.

Mutation, absence or abnormal functioning of retinoblastoma protein are fundamental elements of uncontrolled growth in human cancer. In this study, we analyze the expression of retinoblastoma protein and phosphorylated retinoblastoma protein in melanocytic tumors in vivo. Real-time RT-PCR and immunohistochemistry (tissue microarrays and conventional histological sections) reveal that retinoblastoma protein is progressively upregulated in advanced and metastatic malignant melanomas in vivo. However, this increase is paralleled by increased retinoblastoma protein inactivation due to protein phosphorylation. Interestingly, retinoblastoma protein phosphorylation occurs not homogeneously, but with a 'growth zone'-related pattern. In superficial spreading melanomas a subepidermal-lateral maximum of phosphorylated retinoblastoma protein can be frequently observed. Accordingly, nodular vertically invasive melanomas are characterized by a strong staining of phosphorylated retinoblastoma protein in deep-dermal invading protrusions of the tumor. Furthermore, Kaplan-Meier analysis of 13 cases of advanced melanomas with long-time follow-up suggests a significant negative impact of retinoblastoma protein phosphorylation on survival of melanoma patients independent of tumor thickness. We conclude that the evaluation of phosphorylated retinoblastoma protein in melanocytic tumors could become a helpful adjunct in clinicopathological routine.     

Cytogenetic and molecular investigation of a balanced Xq13q translocation in a patient with retinoblastoma.

We report on a 4-year-old girl with retinoblastoma and de novo balanced translocation [46,X,t (X;13) (q23;q13)]. Unilateral retinoblastoma was discovered at age 9 months along with developmental delay and several manifestations of Turner syndrome. Chromosome studies showed an X/13 translocation and an X inactivation pattern showing the translocated X chromosome active in all 50 cells examined. Standard Southern blot analysis and pulsed field gel electrophoresis using a 3.8 kb EcoR1 fragment of the cDNA probe to the 3' end of the RB1 locus demonstrated a normal genomic pattern. The results of the cytogenetic and molecular analysis suggests that the RB1 locus has not been disrupted by the chromosome rearrangement. This case is the fifth report of an X/13 translocation associated with a retinoblastoma.     

A constitutional mutation within the retinoblastoma gene detected by PFGE

Retinoblastoma may be caused by constitutional mutations in the retinoblastoma gene which segregates as an autosomal dominant inherited predisposition for developing retinoblastoma tumours. Since 75% of these cases are new mutations, there is a need for methods to identify carriers of such germ-line mutations, so that informed genetic counselling is available to patients and close relatives. We have used pulsed-field gel electrophoresis in screening 20 unrelated cases with bilateral retinoblastoma. One constitutional mutation could be detected, and was found to be caused by a balanced chromosome (4; 13) translocation with the breakpoint within intron 17 of the retinoblastoma gene.     

N-myc amplification was rarely detected by fluorescence in situ hybridization in retinoblastoma

In retinoblastoma, genetic alteration of N-myc amplification different from the alteration of the RB1 gene on chromosome 13q14 has been described. This study is to determine the frequency of N-myc amplification by fluorescence in situ hybridization method in retinoblastoma. This study was prospectively derived from 26 patients who were diagnosed as having unilateral retinoblastoma (highly progressive large retinoblastoma, group 5 in Reese-Ellsworth classification) and underwent enucleation. We performed locus-specific fluorescence in situ hybridization probes for N-myc gene. Our results demonstrated that in only one of 26 patients was N-myc amplification found in retinoblastoma tissue. N-myc amplification has been regarded as one characteristic of retinoblastoma cell line and an adverse prognostic factor. However, our study indicates that N-myc amplification is not frequently found in retinoblastoma.     

A deleted chromosome no. 13 in human retinoblastoma cells: relevance to tumorigenesis

In this report of banded karyotypes prepared after short-term culture (72 hr) from human retinoblastoma tumor tissue, one del(13)(pter→q14:) chromosome and one normal chromosome #13 were found in all of the metaphases examined. Similar deletions (always involving 13q14) have previously been described in the somatic cells of individuals with one form of retinoblastoma. In the present case, however, the constitutional karyotype is normal. The presence of tumors in both eyes suggests that this is the genetic form of retinoblastoma, even though the patient's family history is negative for this tumor. The normal constitutional karyotype argues that the chromosome deletion occurred as a postzygotic event. The modal chromosome number of the tumor cells is 47 and rearrangements involving chromosomes #2, #17, and #20 were also identified.     

Family studies on the chromosomal location of the retinoblastoma gene (Rb-1).

The segregation of chromosomes 13 distinguishable by Q band fluorescent polymorphisms has been studied in three families with retinoblastoma. The recombination fraction for two of these families and four families previously reported did not differ significantly from 50%. Since a high recombination fraction has been predicted from chiasma frequency between the centromere of chromosome 13 and 13q14 these results neither confirm nor refute the location of the autosomal dominant gene predisposing to retinoblastoma in 13q14. The use of fluorescent markers is not suitable for early recognition of gene carriers in families with retinoblastoma.     

Retinoblastoma and Hirschsprung disease in a patient with interstitial deletion of chromosome 13

Retinoblastoma is a rare pediatric malignancy (1/20,000) while Hirschsprung disease is a relatively common pediatric disorder (1/5,000). We describe a boy with bilateral retinoblastoma, Hirschsprung disease, multiple minor anomalies, and an interstitial deletion 13q (q13 → q22). This child and a similar previously reported girl with retinoblastoma and Hirschsprung disease may represent a previously unrecognized contiguous gene syndrome. Am. J. Med. Genet. 77:285–288, 1998. © 1998 Wiley-Liss, Inc.     

Location of the retinoblastoma susceptibility gene(s) and the human esterase D locus.

Retinoblastoma occurs with increased frequency in children born with a deletion of the long arm of chromosome 13. Recent reviews have noted that the region 13q14 is consistently deleted in documented cases. Prometaphase and late prophase banding allowed Yunis and Ramsay to determine that a deletion in one patient included the sub-bands q14 . 12, q14 . 13, and q14 . 2, and a portion of q14 . 11 and q14 . 3. We report the results of similar cytogenetic techniques applied in the case of a 26 month old Caucasian female with unilateral retinoblastoma, moderate developmental delay, and subtle dysmorphology. Prometaphase banding of cultured skin fibroblasts revealed the karyotype: mos46,XX/46,XX,del(13)(q13 . 1q14 . 11). Only the sub-band q14 . 11 is deleted in both our patient and that of Yunis and Ramsay. The results are consistent with the localisation of the retinoblastoma susceptibility gene(s) in the sub-band 13q14 . 11. Electrophoretic analysis and activity assays of red blood cell esterase D are consistent with hemizygous expression of that marker in our proband. Comparison with published esterase D analyses in families with retinoblastoma permits the assignment of the esterase D locus to that same sub-band, 13q14 . 11.     

Multiple karyotypic changes in retinoblastoma tumor cells: Presence of normal chromosome No. 13 in most tumors

There are conflicting reports on the frequency in retinoblastoma tumor cells of aberrations involving chromosome No. 13. To quantitate the frequency of various chromosome aberrations, we analyzed the karyotypes from ten retinoblastoma tumors; all tumors contained chromosome abnormalities. Chromosome No. 13 was altered in only two tumors, but the aberrations in these two cases affected different portions of the chromosome. We have concluded that chromosome aberrations affecting chromosome No. 13 are relatively infrequent in retinoblastoma tumors. Chromosome No. 1 was involved in rearrangements in eight tumors; in six tumors the rearrangements lead to trisomy of 1q25-1q32. Seven tumors had aberrations resulting in trisomy of the long arm of chromosome No. 17; the most common aberration was an i(17q) chromosome. Every tumor showed trisomy of the long arm of either chromosome No. 1 or 17. These changes in chromosomes No. 1 and 17 have been observed by others in many different tumors and are not unique to retinoblastoma. In summary, chromosome abnormalities were present in all retinoblastoma tumors studied, but no aberration common to all tumors was found.     

Chromosome abnormalities involving band 13q14 in hematologic malignancies

Fifteen patients with hematologic disorders showed abnormalities involving chromosome band 13q14. Nine patients had an interstitial deletion of this band, similar to that reported in some retinoblastoma tumors and as a constitutional abnormality in a small proportion of cases of familial retinoblastoma. In five patients, band 13q14 was involved in translocations and in one case there was a deletion of one chromosome #13 and a translocation involving the homologous #13. The diagnosis in the majority of our patients (11 of 15) was chronic lymphocytic leukemia. In these patients the abnormalities were detected in cultures stimulated with 4-phorbol 12-myristate 13-acetate (PMA). It is possible that the utilization of this agent is a fundamental requirement for the reliable demonstration of abnormalities involving 13q14 in patients with B-cell malignancies. The incidence of abnormalities involving 13q14 and their significance in the development of neoplasias, other than retino-blastoma, is discussed.     

Isolation of human chromosome 13-specific DNA sequences cloned from flow sorted chromosomes and potentially linked to the retinoblastoma locus

A recombinant DNA library has been constructed using flow sorted chromosome #13 DNA and the phage vector, Charon 21A. Roughly 90% of the phage inserts in the library hybridize to human repetitive DNA. Phage containing human nonrepetitive inserts have been screened for chromosome #13 specificity by Southern blot analysis using the genomic DNA of human-rodent cell hybrids containing different regions of the human #13 autosome. Of 18 phage inserts characterized, 13 have been assigned to the 13q12----q22 subregion, three appear to be localized in the 13pter----q12 region, and two are not #13-specific. By Southern blot analysis of the DNA of a retinoblastoma patient exhibiting a deletion of band 13q14 and of karyotypically normal individuals, two phage inserts have been putatively assigned to band 13q14, the currently accepted locus for a genetic determinant for retinoblastoma. These two DNA probes show quantitative differences in hybridization band intensity in the genomic DNA of the 13q--patient relative to that of the normals. In situ hybridization data support these conclusions. A recombinant phage library that shows an approximate 90% enrichment for human chromosome #13-specific DNA fragments should prove useful not only in studies related to retinoblastoma, but also in the molecular analysis of the structure and function of chromosome #13.