Unexplained developmental delay/learning disability: guidelines for best practice protocol for first line assessment and genetic/metabolic/radiological investigations.

BACKGROUND: Investigation of patients, particularly children, with unexplained global developmental delay (GDD)/learning disability (LD) has been challenging due to a lack of clear guidance from specialised centres. Limited knowledge of rare diseases and a poor understanding of the purpose or limitations of appropriate investigations have been some of the principal reasons for this difficulty. AIMS: A guideline development group was formed to recommend on appropriate, first line metabolic, genetic and radiological investigations for children and adults with unexplained GDD/ID. METHODS AND RECOMMENDATIONS: A comprehensive literature search was conducted, evaluated and reviewed by the guideline committee and a best practice protocol for first line assessment and genetic, metabolic and radiological investigations was decided upon after considering diagnostic yield, practicality, treatability and costs. CONCLUSION: It is hoped that these recommendations will become national guidelines for the first line metabolic, genetic and radiological investigation of patients presenting with unexplained GDD/ID.

Prenatal origin of separate evolution of leukemia in identical twins.

Several studies involving identical twins with concordant leukemia and retrospective scrutiny of archived neonatal blood spots have shown that the TEL-AML1 fusion gene in childhood acute lymphoblastic leukemia (ALL) frequently arises before birth. A prenatal origin of childhood leukemia was further supported by the detection of clonotypic immunoglobulin gene rearrangements on neonatal blood spots of children with various other subtypes of ALL. However, no comprehensive study is available linking these clonotypic events. We describe a pair of 5-year-old monozygotic twins with concordant TEL-AML1-positive ALL. Separate leukemic clones were identified in the diagnostic samples since distinct IGH and IGK-Kde gene rearrangements could be detected. Additional differences characterizing the leukemic clones included an aberration of the second, nonrearranged TEL allele observed in one twin only. Interestingly, both the identical TEL-AML1 fusion sequence and distinct immunoglobulin gene rearrangements were identified on the neonatal blood spots indicating that separate preleukemic clones evolved already before birth. Finally, we compared the reported twins with an additional 31 children with ALL by using the microarray technology. Gene expression profiling provided evidence that leukemia in twins harbours the same subtype-typical feature as TEL-AML1-positive leukemia in singletons suggesting that the leukemogenesis model might also be applicable generally.

Abnormalities of chromosome 16q in myeloid malignancy: 14 new cases and a review of the literature.

Fourteen patients with abnormalities of chromosome 16q, 13 with acute myelogenous leukaemia (AML), and one with refractory anaemia with excess of blasts (RAEB), are described. Seven patients had inv(16)(p13q22), two had del(16)(q22), and five had other abnormalities of 16q. Six of the seven patients with inv(16) had AML M4Eo and, following treatment with adriamycin, cytosine arabinoside, and 6-thioguanine, all achieved complete remission (CR). Neither patient with del(16)(q22) had typical M4Eo morphology at diagnosis; CR was achieved in one and one had resistant leukaemia. Patients with other abnormalities of 16q had blasts of diverse morphology and, although morphologically abnormal eosinophils were seen in three patients, this was not as marked as in the patients with inv(16). CR was achieved in two of the four patients with other abnormalities of 16q but duration of remission was short in both cases. These results suggest that most patients with del(16)(q22) and other abnormalities of 16q22 do not have typical AML M4Eo. Such patients tend to have a worse prognosis, and are more likely to have complex karyotypes typical of secondary leukaemia.

Cytogenetics in acute myeloid leukaemia.

A wealth of literature spanning 20 years describing cytogenetic abnormalities in acute myeloid leukaemia (AML) already exists. It ranges from single case reports of unusual abnormalities to large multicentre studies of hundreds of cases. A landmark publication was the Fourth International Workshop on Chromosomes in Acute Leukaemia which established a base line for diagnosis, prognosis and frequency of chromosome abnormalities in AML. Two large sources of information are a book, 'The Chromosomes in Human Cancer and Leukemia' and a catalogue of chromosome abnormalities, which aims to list all chromosome abnormalities described in the scientific and medical literature from 1973, when the widespread use of banding techniques, enabled the precise definition of the chromosome breakpoints. In this review the common cytogenetic abnormalities seen in AML with reference to associations with the French-American-British (FAB) classification, their possible prognostic significance and their associated molecular biology are summarized.

Severe hypodiploidy with karyotype instability in a case of acute myeloid leukemia.

We present a case of acute myeloid leukemia (AML) in an elderly male. A severe hypodiploid karyotype (chromosome range 29-39) was detected with a large amount of cell-to-cell variation, suggesting that the leukemic cells are primarily characterized by chromosomal loss due to karyotypic instability. Severe hypodiploidy is a rare finding in AML but previous similar cases indicate that it confers a very poor prognosis.

Cytogenetic analysis in a case of intraocular medulloepithelioma.

We report the cytogenetic analysis of a medulloepithelioma, a rare neuroectodermally-derived tumor of childhood, which to our knowledge is the first reported karyotype of this tumor. The tumor sample was received at diagnosis and was mechanically dissociated to create cell suspension cultures. The primary cytogenetic abnormality was a der(16)t(1;16) chromosome, which was typical of that reported in a wide range of other tumor types. The other abnormalities seen were a del(6q) and monosomy 15.

Routine karyotyping in Wilms tumor.

We describe the karyotypes of nine Wilms tumors (WT). Four tumors were initially karyotyped from diagnostic needle core biopsies, 3 after postchemotherapy tumor resection and the remainder from xenografts grown in nude mice. The 9 nephroblastomas were composed of 7 with favorable histology (intermediate-grade malignancy) and 2 with unfavorable histology (anaplastic or high-grade malignancy). The 7 tumors with favorable histology had karyotypes typical of WT, with the previously described nonrandom abnormalities +1q, +6, +7, +8, +12, +13, +18 and structural abnormalities of 1p and 16q present in at least 1 case. The most common abnormalities were trisomy 18 (4 cases) and +1q (3 cases). The 2 tumors with unfavorable histology both had complex karyotypes atypical for WT. We suggest that cytogenetics can act as a marker when histologic grade is in doubt. Karyotypic analysis from needle core biopsies was attempted in 6 samples, including 1 from a nephrogenic rest (NR) of the nonaffected kidney and provided a result on 5 occasions. The NR were present in the sole case with a constitutional abnormality, a mosaic partial duplication of 8q. However, both the tumor and the NR were apparently derived from the normal cell line. Here we demonstrate that a cytogenetic result can be routinely obtained from needle core biopsies and will thus facilitate true diagnostic tumor karyotypes in both WT and other tumors.

Loss of X chromosome in childhood acute lymphoblastic leukemia.

We present six cases of childhood acute lymphoblastic leukemia (ALL) in which an acquired loss of the X chromosome was detected. The cases derive from a consecutive series of 178 childhood ALL, consisting of 80 girls and 98 boys. In five cases the presence of the TEL-AML1, t(12;21), fusion product was detected by FISH. The single negative case had an unusual t(1;19)(p13;q13). In addition, this was the only case that did not have a cytogenetically visible rearrangement involving one of the chromosome regions 6q, 9p, or 12p. The six cases showed the typical presentation features of an ALL of FAB type L1, a common ALL immunophenotype with myeloid marker co-expression, and a median presenting age of 7 years. We, therefore, conclude that loss of chromosome X may be a secondary event in older girls with TEL-AML1-positive ALL.

Trisomy 1q generating translocations in Wilms tumor.

Unbalanced translocations generating trisomy of 1q are common in Wilms tumor (WT). We present eight unbalanced 1q translocations from seven tumors and a review of the literature. An unbalanced translocation that results in a der(16)t(1q;16q) chromosome represents more than half of the published +1q generating translocations in WT. This translocation is also common to many other tumor types. Four of the tumors presented here contained this chromosome and,in two cases, it was the primary acquired cytogenetic abnormality within the tumor. The other four translocations involved 9q31, 9q34, 17p1?, and 21p11 as the partner to 1q. The chromosome 17 and 21 translocations occurred within the same tumor as apparently independent events. In contrast with the 16q translocations, these other translocations were secondary cytogenetic events, thereby indicating a role in tumor progression rather than initiation. Probes mapping to 1q12 and 1q21 were employed for fluorescence in situ hybridization and it was demonstrated that different 1q breakpoints are possible. In this series, the majority of breakpoints either mapped to 1q12 or were centromeric to this region.

The investigation of karyotypic instability in the high-hyperdiploidy subgroup of acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) of childhood has been cytogenetically well characterized, and approximately 25% of cases will have a high-hyperdiploid (51-68) chromosome complement. In a 5 year period a consecutive series of 152 presentation ALL's were karyotyped. In all cases a result was obtained and 138 (91%) had a detectable abnormal clone of which 44 (29%) were high-hyperdiploid. Within the high-hyperdiploidy group karyotypic cell to cell variation was observed in many cases. To provide further evidence of this phenomenon a dual-color fluorescence in-situ hybridization (FISH) experiment was performed on stored fixed suspension from 14 ALL's with such a karyotype. In each case 4-6 probes were investigated, employing probes to centromeres of chromosomes X, 4, 6, 8, and 10 and a locus specific probe to chromosome 21q22. It was found that the FISH produced results that were generally in good agreement with the G-banding findings and supported the notion of karyotypic cell to cell variation. FISH further showed that most of cases would have two extra copies of chromosome 21 in the majority of leukemic cells and a single extra copy in the minority. A further finding was that fewer cells contained extra copies of chromosomes 6, 8 and 10 than was expected based on the comparison of the signal number of the other probes investigated. In contrast chromosomes X, 4, and 21 seldom displayed this feature. We have demonstrated that karyotypic instability as defined by karyotypic cell to cell variation is a feature of the high-hyperdiploid subgroup in childhood ALL. It is questioned whether the underlying defect resulting in the observed karyotypic instability of this subgroup is one of the primary causative events in the formation of the leukemia.

Primary pleomorphic adenoma of the external auditory canal diagnosed by fine needle aspiration cytology. A case report.

BACKGROUND: Pleomorphic adenoma (PA) arising in the external auditory canal (EAC) is a very rare neoplasm, thought to be derived from ceruminous glands. CASE: A 43-year-old male presented with a slowly growing mass in the right EAC. Clinical and radiologic examinations showed a well-circumscribed tumor limited to the EAC, without a connection to the parotid gland. Fine needle aspiration cytology (FNAC) revealed the typical cytologic findings of PA. The diagnosis was confirmed by histologic examination. CONCLUSION: This case illustrates that together with clinical and radiologic findings, primary PA of the EAC can confidently be diagnosed by FNAC.

Duplication of 17q11.2 and Features of Albright Hereditary Osteodystrophy Secondary to Methylation Defects within the GNAS Cluster: Coincidence or Causal?

We report a case of Albright hereditary osteodystrophy (AHO) in a three-year-old girl with a microduplication at 17q11.2. The child developed obesity within the first 6 months of life. A diagnosis of Albright was made at age 2 years when biochemical evidence of parathyroid resistance was found. No mutations were identified in guanine nucleotide-binding protein G (s) subunit alpha (GNAS1). Subsequent investigations revealed methylation disturbance at GNAS1A, neuroendocrine secretory protein antisense (NESPAS) and neuroendocrine secretory protein 55 (NESP55) confirming a diagnosis of pseudohypothyroidism type 1B. A deletion of NESP55 and uniparental disomy chromosome 20 were excluded which suggested that the features of AHO arose through a purely epigenetic mechanism. Further investigation revealed a de novo microduplication at 17q11.2 encompassing the neurofibromatosis type 1 (NF1) gene. The combination of two rare de novo events in the same child raises the possibility that duplication of a gene within the 17q11.2 region may have triggered abnormal methylation in the GNAS cluster region on chromosome 20.

Deletion 14q (q22q23) associated with anophthalmia, absent pituitary, and other abnormalities.

A fetus is described with anophthalmia, absent pituitary, hypoplastic adrenal glands and kidneys, absent left horn of the uterus, underdeveloped genitalia, and clinodactyly, with a deletion of 14(q22q23). A review of published reports found no similar deletion cases.

A 45,X/69,XXY fetus.

Post-mortem examination of a 20-week fetus showed incompletely masculinised external genitalia, hypoplastic adrenal glands and minor physical stigmata suggestive of a chromosome abnormality. Gonad and skin were karyotyped and both were found to contain two cell lines, 45,X and 69,XXY. It appears this fetus is a true 45,X/69,XXY mosaic.

Constitutional balanced chromosomal rearrangements and neoplasm in children.

A predisposition to tumor development is currently associated with some, but not all, constitutional chromosomal abnormalities. In a series of 578 children, in which conventional cytogenetic investigation was performed on material from various benign and malignant tumors, four boys and one girl were also found to have constitutional balanced chromosomal rearrangements. The figure of 5 in 578 is notable because the reported incidence of balanced rearrangements in newborns is approximately 1 in 450. Thereby suggesting that some, if not all, children with balanced constitutional chromosomal rearrangements have an increased predisposition for neoplasms developing.

A further case of a t(11;20)(p15;q11.2) translocation in an acute myeloid leukemia (FAB M2).

PURPOSE: The clinical implications of many cytogenetic abnormalities in acute myeloid leukemia (AML) are now well established. However, questions about the significance of rarer abnormalities still exist, particularly in childhood disease. PATIENTS AND METHODS: We report a case of a 9 1/2-year-old girl who had AML of the FAB M2 subtype. A diagnostic bone marrow aspirate and subsequent bone marrow aspirates were investigated using conventional cytogenetic methods. RESULTS: Cytogenetic analysis of the diagnostic bone marrow aspirate showed a t(1;20)(p15;q11.2) translocation as the sole acquired abnormality. After one course of chemotherapy, the patient achieved hematopoietic and cytogenetic remission which has been sustained for 1 year after presentation. CONCLUSION: This report demonstrates that rare non-random cytogenetic abnormalities are still to be described in AML, and that care should be taken when ascribing clinical significance to cytogenetic findings in childhood disease based on those of older patients.

Monosomy 7, diabetes insipidus and acute myeloid leukemia in childhood.

The triad of diabetes insipidus (DI), monosomy 7, and acute myeloid leukemia in a 7-year-old boy is described. This triad has been described in adults but not in children. The DI ran a transient, self-limiting course and required no specific therapy. The pathogenesis of DI remains unknown, and its transient nature may result in this component of the triad going unnoticed.

Molecular features of a human rhabdomyosarcoma cell line with spontaneous metastatic progression.

A novel human cell line was established from a primary botryoid rhabdomyosarcoma. Reverse transcription polymerase chain reaction investigations of this cell line, called RUCH-2, demonstrated expression of the regulatory factors PAX3, Myf3 and Myf5. After 3.5 months in culture, cells underwent a crisis after which Myf3 and Myf5 could no longer be detected, whereas PAX3 expression remained constant over the entire period. Karyotype analysis revealed breakpoints in regions similar to previously described alterations in primary rhabdomyosarcoma tumour samples. Interestingly, cells progressed to a metastatic phenotype, as observed by enhanced invasiveness in vitro and tumour growth in nude mice in vivo. On the molecular level, microarray analysis before and after progression identified extensive changes in the composition of the extracellular matrix. As expected, down-regulation of tissue inhibitors of metalloproteinases and up-regulation of matrix metalloproteinases were observed. Extensive down-regulation of several death receptors of the tumour necrosis factor family suggests that these cells might have an altered response to appropriate apoptotic stimuli. The RUCH-2 cell line represents a cellular model to study multistep tumorigenesis in human rhabdomyosarcoma, allowing molecular comparison of tumorigenic versus metastatic cancer cells.