Parental occupational exposure and the risk of acute lymphoblastic leukemia in offspring in Israel.

OBJECTIVE: Parental employment in occupations that have potential exposures to organic solvents or pesticides could be associated with the risk of childhood acute lymphoblastic leukemia (ALL) in their offspring. METHODS: We explored this hypothesis by studying the association with respect to exposure time windows. Our case-control study included 224 children, 112 diagnosed with ALL and 112 matched controls. RESULTS: A significantly higher odds ratio (OR) was found between childhood ALL and reported parental occupational exposures. Analysis of exposures of both parents by exposure time windows revealed significant OR during the preconception and postnatal periods separately. CONCLUSIONS: The results provide support to the association between parental occupational exposures and ALL in their children. These results should be interpreted cautiously because of the small numbers, biases characterizing case-control studies, and the use of hospital-based controls.

Prediction of high risk Ewing's sarcoma by gene expression profiling.

Ewing's sarcoma (ES) is the second most common primary malignant bone tumor in children and adolescents. Currently accepted clinical prognostic factors fail to classify ES patients' risk to relapse at diagnosis. We aimed to find a new strategy to distinguish between poor and good prognosis ES patients already at diagnosis. We analysed the gene expression profiles of 14 primary tumor specimens and six metastases from ES patients, using oligonucleotide microarray analysis. The over-expression of two genes was validated by quantitative PCR using the LightCycler system. We identified two distinct gene expression signatures distinguishing high-risk ES patients that are likely to progress from low-risk ES patients with a favorable prognosis of long-term progression-free survival. The microarray-based classification was superior to currently used prognostic parameters. Over-expressed genes in the poor prognosis patients included genes regulating the cell cycle and genes associated with invasion and metastasis, while among the downregulated genes were tumor suppressor genes and inducers of apoptosis. Our results indicate the existence of a specific gene expression signature of outcome in ES already at diagnosis, and provide a strategy to select patients who would benefit from risk-adapted improved therapy.

CATSPER2, a human autosomal nonsyndromic male infertility gene.

In the course of positional cloning of the Congenital Dyserythropoietic Anemia type I (CDAI) [MIM 224120] gene on 15q15.1-15.3, we examined a family of French origin, in which the propositus suffered from asthenoteratozoospermia and nonsyndromic deafness in addition to CDAI. Two of his brothers had a similar phenotype. All three siblings were homozygous carriers of the CDA1 mutation as well as of a distally located approximately 70 kb deletion of the proximal copy of a 106 kb tandem repeat on chromosome 15q15. These repeats encode four genes whose distal copies may be considered pseudogenes. Lack of functional stereocilin and CATSPER2 (a voltage-gate cation channel expressed specifically in spermatozoa) may explain the observed deafness and male infertility phenotypes. To the best of our knowledge, the involvement of CATSPER2 in asthenoteratozoospermia is the first description of a human autosomal gene defect associated with nonsyndromic male infertility.

der(11)t(11;17): a distinct cytogenetic pathway of advanced stage neuroblastoma (NBL) - detected by spectral karyotyping (SKY).

Conventional cytogenetic, molecular cytogenic and genetic methods disclosed a broad spectrum of genetic abnormalities leading to gain and loss of chromosomal segments in advanced stage neuroblastoma (NBL). Specific correlation between the genetic findings could delineate distinct genetic pathways, of which the biology and prognostic significance is as yet undetermined. Using spectral karyotyping (SKY) and fluorescence in situ hybridization (FISH) on metaphases from 16 patients with advanced stage NBL, it was possible to explore the whole spectrum of rearrangement within complex karyotypes and to detect hidden recurrent translocations. All translocations were unbalanced. The most prevalent recurrent unbalanced translocations resulted in 17q gain in 12 patients (75%), 11q loss in nine patients (56%), and 1p deletion/imbalance in eight patients (50%). The most frequent recurrent translocation was der(11)t(11;17) in six patients. Three cytogenetic pathways could be delineated. The first, with six patients, was characterized by the unbalanced translocation der(11)t(11;17), detected only by SKY, resulting in the concomitant 17q gain and 11q loss. No MYCN amplification or 1p deletion (except one patient with 1p imbalance) were found, while 3p deletion, and complex karyotypes were common. The second subgroup, with four patients, had 17q gain and 1p deletion, and in two patients 11q loss, that was apparent only by FISH. 1p deletion occurred through der(1)t(1;17) or del(1p). The third subgroup of four patients was characterized by MYCN amplification with 17q gain and 1p deletion, very rarely with 11q loss (one patient) through a translocation with a non-17q partner. The SKY subclassifications were in accordance with the findings reported by molecular genetic techniques, and may indicate that distinct oncogenes and suppressor genes are involved in the der(11)t(11;17) pathway of advanced stage NBL.

Ten novel Diamond-Blackfan anemia mutations and three polymorphisms within the rps19 gene.

INTRODUCTION: A total of 25% of patients presenting with Diamond-Blackfan anemia (DBA) carry mutations in the rps19 gene, which encodes protein RPS19 of the small ribosomal subunit. The other DBA cases carry mutations in other, unknown gene(s). MATERIALS AND METHODS: We searched mutations in 48 DBA families or isolated patients based on PCR of exons of the rps19 gene and automatic sequencing. We also studied three novel intronic polymorphisms in 85 persons (most of the patients and their relatives, when the latter could be investigated). RESULTS: We identified 10 new mutations within the rps19 gene. We found no obvious correlation between the clinical expression and the nature of the mutation. Besides, we found three polymorphisms within the rps19 gene. Polymorphisms a, b and c were (i) a one-base insertion (+c at position +79) in intron 2, (ii) the g-->c substitution at position +89 also in intron 2 and (iii) the g-->a substitution at position +14 in intron 4. Inheritance studies showed that the polymorphisms were transmitted en bloc, thus defining the --- haplotypes (changes absent) and the +++ haplotypes (changes present). The percentages of each haplotype were about 50% in families and isolated persons with DBA, as well as in controls. CONCLUSION: For the 10 novel mutations found in the rps19 gene, there were no obvious genotype-phenotype correlations. The transmission of the polymorphisms was en bloc and the studies did not suggest any clinical correlates at this stage.

ATM gene mutations are not involved in medulloblastoma in children.

Primitive neuroectodermal tumors (PNET)-medulloblastomas account for approximately 20% of all brain tumors in children. Ataxia-telangiectasia is an autosomal recessive neurological disorder with predisposition to cancer. The most common neoplasms are lymphoid malignancies and solid tumors, including central nervous system tumors, astrocytomas, and medulloblastomas. To investigate the potential role of the ATM gene in the pathogenesis of medulloblastoma, 13 tumors were screened for ATM mutations and 9 for loss of heterozygosity (LOH) of the ATM locus and flanking regions. In none of the tumors were mutations identified. In five of them, the well-known polymorphisms D1853N and F858L were identified and in all 22 tumors, the wild-type allele was preserved. The frequency of the polymorphisms was similar to that reported in our and other normal populations. The LOH of the 11q region (including the ATM gene), detected in 25% of informative cases, is consistent with the molecular and cytogenetic reports of deletion of chromosome 11 in 13%-41% of medulloblastomas. These results indicate that mutations in the ATM gene do not play a role in the pathogenesis of medulloblastoma in children. The LOH in the 11q region may suggest hidden unidentified tumor suppressor genes that may be involved in the malignant transformation.

High frequency of genomic instability in Ewing family of tumors.

We tested Ewing sarcoma tumors for microsatellite instability (MSI) and loss of heterozygosity (LOH) to investigate the role of genomic instability (GI) in this sarcoma. We detected a high frequency of GI (57%), mostly on 1p and 11p, 35% and 30%, respectively. Patients with GI compared to those with stable genome had a median progression-free survival (PFS) and overall survival (OS) of 24 months and 70 months, compared with 39 and 84 months, respectively. MSI was observed in 48% (11/23) of the tumor samples. Low-MSI (L-MSI) patients (with MSI presented at only one locus) tended to have a better prognosis, 70% PFS, compared with 25% in the high-MSI (H-MSI) group (P=0.13). LOH without MSI did not correlate with progression. H-GI (MSI and/or LOH in > or =30% of tested markers) tended to associate with an adverse prognosis (P=0.28), and correlated significantly with the pelvic site of the primary tumor (P=0.02). The instability of 1p was not associated with progression, while alterations at the 11p locus tended to correlate with a more aggressive disease (P=0.18). Our data suggest that GI may play a role in Ewing sarcoma clinical behavior and outcome.

Telomere length is a prognostic factor in neuroblastoma.

BACKGROUND: Maintenance of telomeres, in most instances by reactivation of telomerase, is obligatory for the indefinite proliferation of tumor cells. The objective of this study was to evaluate telomere length and telomerase activity (TA) as markers for progression and prognosis in neuroblastoma. METHODS: Primary tumor samples from 51 patients were analyzed for telomere length and TA and were correlated with known prognostic parameters and outcome. RESULTS: Telomere length had a highly significant correlation with prognosis (P = .007). Short telomeres were predictive of a favorable prognosis, whereas long or unchanged telomeres were predictive of a poor outcome. For the first time to their knowledge, the authors have shown that, within the high-risk group patients, telomere length could define a favorable subgroup that had a progression-free survival (PFS) rate of 86% compared with a PFS rate of 36% for patients with more adverse disease, which is the expected PFS rate for such patients (P = .04). In a multivariate analysis, telomere length was the most significant prognostic parameter (P = .032). TA was correlated significantly with outcome and with known prognostic factors. High TA and low TA were associated with adverse and favorable outcomes, respectively (P = .01). CONCLUSION: The results of this investigation suggested that telomere length is a highly significant prognostic parameter of clinical relevance in patients with neuroblastoma. In high-risk patients, telomere length was the sole significant parameter that identified a group of patients who had a favorable prognosis. The authors suggest that telomere length should be included in the recommended diagnostic investigations for patients with neuroblastoma.

Acquired trisomy 21 and distinct clonal evolution in acute megakaryoblastic leukaemia in young monozygotic twins.

An intrauterine origin of childhood acute lymphoblastic leukaemia (ALL) was proven by the identical clonotypic gene rearrangement in the concordant leukaemias of monozygotic twins, arising from a single clonogenic progeny. The monozygotic twins, presented at the age of 22 months with acute megakaryoblastic leukaemia (AML-M7) in one and myelodysplasia transformed to AML-M7 in the other. Leukaemic cells in both twins carried trisomy 21 and additional different clonal evolution changes of del(20q) in the first twin and trisomy 8 in the second. AML-M7 of late infancy with trisomy 21 may be included in the leukaemias of intrauterine origin, possibly a result of genotoxic insult.

The predictive potential of molecular detection in the nonmetastatic Ewing family of tumors.

BACKGROUND: Tumors in the Ewing family (EFTs) are the second most common bone tumors in children and adolescents. Despite aggressive chemotherapy, one-third of patients with localized tumor still may develop recurrences. This implies that not all tumor cells are eradicated and that the patients may have a level of residual disease. EFTs are characterized by specific chromosomal translocations that result in chimeric transcripts that can be detected with reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. METHODS: The authors report the prognostic potential of the positive chimeric transcript (EWS/FLI1) in bone marrow (BM) and/or peripheral blood (PBL) in 26 patients with EFT during a long follow-up period (median, 61 months). RESULTS: At diagnosis, 43% of patients had positive RT-PCR BM results, with no correlation to tumor progression (P = 0.3). During follow-up, 58% of patients had positive RT-PCR results in their last sample analyzed (BM and/or PBL). A highly significant correlation between the presence of the chimeric transcript and disease progression was detected (P = 0.0028). In a multivariate analysis, the percentage of tumor necrosis (P = 0.007) and RT-PCR results during follow-up (P = 0.02) remained significant prognostic markers. In 10 of 11 patients who developed disease progression, BM and/or PBL samples were positive for the chimeric transcript before evidence of overt clinical recurrence. CONCLUSIONS: Occult tumor cells in BM and/or PBL samples during long follow-up are strong predictors of recurrent disease in patients with nonmetastatic EFTs.

Germ-line ATM gene alterations are associated with susceptibility to sporadic T-cell acute lymphoblastic leukemia in children.

A major feature of ataxia-telangiectasia (A-T) is an increased risk of cancer, particularly of lymphoid malignancies. We studied ATM gene involvement in leukemic cells derived from 39 pediatric T-cell acute lymphoblastic leukemias (ALLs). Two types of sequence changes--truncating and missense--were identified in 8 T-cell ALL samples: 3 truncating changes, all previously identified in A-T (R35X, -30del215, 2284delCT), and 3 missense variants (V410A, F582L, F1463C) were found, none associated with loss of heterozygosity (LOH). In all patients studied, the mutation was present in the germ-line. A-T carriers, defined by the finding of truncating mutations, were found to be 12.9 times more frequent than in the normal population (P = 0.004). A normally ethnically matched population was screened for the 3 missense variants, and their frequency was significantly more prevalent (4.9-fold excess) than in the normal population (P = 0.03). Our data suggest there is some evidence of an association between missense alterations in the ATM gene and T-cell ALL. A significant difference in the mean age at diagnosis of T-cell ALL was noted between patients harboring an ATM sequence change and those with no change, 5.4 years and 9.7 years, respectively (P = 0.001). No ATM alterations were identified in relapse samples, indicating that ATM does not play a role in disease progression. The high prevalence of germ-line truncating and missense ATM gene alterations among children with sporadic T-cell ALL suggests an association with susceptibility to T-cell acute leukemia and supports the model of predisposition to cancer in A-T heterozygotes.

Prenatal diagnosis in Li-Fraumeni syndrome.

PURPOSE: The hallmark of Li-Fraumeni syndrome (LFS), a familial cancer syndrome, is constitutional TP53 mutation. The authors addressed the complex question of predictive prenatal genetic testing for cancer risk associated with inheritance of TP53 mutation. METHODS: A classic LFS family including the proband (a 20-month-old boy with rhabdomyosarcoma), his 36-year-old father with osteosarcoma, and his 40-year-old paternal aunt with bilateral breast cancer were identified as carriers of a TP53 germline mutation, a novel 1 base pair deletion in exon 5. A few years later, the mother became pregnant twice, and the parents requested prenatal diagnosis on each occasion. Genetic counseling, psychological evaluation, and support were provided by a multidisciplinary team including a pediatric oncologist, a geneticist, a psychosocial worker, a prenatal care provider, and an ethical representative. After providing overall information on LFS, including the high risk of developing secondary multiple neoplasms in LFS survivors, the committee approved prenatal diagnosis at the request of the family. RESULTS: In the two pregnancies, the two fetuses were found to be carriers of the same mutation. Nine years from diagnosis of the first tumor, the proband, and a month later his father, developed second tumors, multifocal osteosarcoma and leiomyosarcoma, respectively. CONCLUSIONS: Children with primary tumors belonging to LFS should be considered for screening for germline mutations and genetic counseling by a multidisciplinary team. Whether family members are found to be positive or negative as carriers, such measures may provide, by reducing uncertainty, psychological benefit to high-risk families.

Distinct cytogenetic pathways of advanced-stage neuroblastoma tumors, detected by spectral karyotyping.

Molecular studies of advanced-stage neuroblastoma (NBL) have revealed a marked genetic heterogeneity. In addition to MYCN amplification and chromosome 1 short-arm deletions/translocations detected by conventional cytogenetics, application of fluorescence in situ hybridization has disclosed a high prevalence of 17q gain, whereas allelotyping and comparative genomic hybridization techniques also have revealed loss of 11q and of other chromosomal material. Using the recently developed technique of spectral karyotyping (SKY), we sought to refine the cytogenetic information, identify hidden recurrent structural chromosomal abnormalities, and compare them to the molecular findings. Thirteen samples of metaphase spreads from 11 patients with advanced-stage NBL were analyzed by SKY. Most of them were found to have complex karyotypes (more than three changes per metaphase) and complex unbalanced rearrangements. Recurrent aberrations leading to 17q gain, deletion of 1p, MYCN amplification, and loss of 11q appeared in 7, 4, 4, and 5 patients, respectively, in simple and complex karyotypes. Chromosome 3 changes and gain of 1q and 7q appeared in 6, 5, and 4 patients, respectively, in complex karyotypes only, reflecting later changes. A strikingly high prevalence of the unbalanced translocation der(11)t(11;17), leading to concomitant 11q loss and 17q gain in 4 patients, delineated a distinct cytogenetic group, none having 1p deletion and/or MYCN amplification. der(11)t(11;17) was associated with complex karyotypes with changes in chromosomes 3 and 7q. The 17q translocations with partners other than 11q were associated with 1p deletion and/or MYCN amplification. The distinct cytogenetic subgroups identified by SKY confirm and extend the recent molecular observations, and suggest that different genes may interact in the der(11)t(11;17) pathway of NBL development and progression.

Classical and molecular cytogenetic abnormalities and outcome of childhood acute myeloid leukaemia: report from a referral centre in Israel.

The incidence of cytogenetic abnormalities in childhood de novo acute myeloid leukaemia (AML) and its prognostic significance was assessed in an Israeli paediatric referral centre. Cytogenetic analysis was successful in 86 of 97 children (< 20 years of age) diagnosed between 1988 and 2002 with de novo AML. Fluorescence in situ hybridization analysis detected new information in 11 of them, leading to reassignment in cytogenetic group classification. The incidence of the various cytogenetic subgroups was as follows: normal - 9%; t(11q23) - 22%; t(8;21) - 13%; t(15;17) - 8%; inv(16) - 3.4%; abn(3q) - 4.6%; 7/7q-(sole or main) - 5.8%; del(9q)(sole) and +21(sole) - 4.6% each; t(8;16) - 2.3%; t(6;9), t(1;22), +8(sole) - 1.1% each; and miscellaneous - 18%. The overall survival (OS) and event-free survival (EFS) (4 years) for 94 patients treated with the modified Berlin-Frankfürt-Münster (BFM) AML protocols (non-irradiated) were 59.9% (SE = 5%) and 55.7% (SE = 5%), respectively, and for the favourable t(8;21), t(15;17) and inv(16), OS was 60% (SE = 15%), 83% (SE = 15%) and 100% respectively. For the normal group it was 62% (SE = 17%), miscellaneous 64% (SE = 12%), t(11q23) 44.6% (SE = 11%) and of the -7/7q-, del(9q)(sole) or t(6;9), none had survived at 4 years. The incidence of cytogenetic subgroups in the Israeli childhood AML population and their outcome were similar to other recently reported paediatric series. Cytogenetic abnormalities still carry clinical relevance for treatment stratification in the context of modern chemotherapy.

Congenital dyserythropoietic anemia type I is caused by mutations in codanin-1.

Congenital dyserythropoietic anemias (CDAs) constitute a rare group of inherited red-blood-cell disorders associated with dysplastic changes in late erythroid precursors. CDA type I (CDAI [MIM 224120], gene symbol CDAN1) is characterized by erythroid pathological features such as internuclear chromatin bridges, spongy heterochromatin, and invagination of the nuclear membrane, carrying cytoplasmic organelles into the nucleus. A cluster of 45 highly inbred Israeli Bedouin with CDAI enabled the mapping of the CDAN1 disease gene to a 2-Mb interval, now refined to 1.2 Mb, containing 15 candidate genes on human chromosome 15q15 (Tamary et al. 1998). After the characterization and exclusion of 13 of these genes, we identified the CDAN1 gene through 12 different mutations in 9 families with CDAI. This 28-exon gene, which is transcribed ubiquitously into 4738 nt mRNA, was reconstructed on the basis of gene prediction and homology searches. It encodes codanin-1, a putative o-glycosylated protein of 1,226 amino acids, with no obvious transmembrane domains. Codanin-1 has a 150-residue amino-terminal domain with sequence similarity to collagens and two shorter segments that show weak similarities to the microtubule-associated proteins, MAP1B (neuraxin) and synapsin. These findings, and the cellular phenotype, suggest that codanin-1 may be involved in nuclear envelope integrity, conceivably related to microtubule attachments. The specific mechanisms by which codanin-1 underlies normal erythropoiesis remain to be elucidated.