Prognostic Impact of Node-Spreading Pattern in Surgically Treated Small-Cell Lung Cancer: A Multicentric Analysis.

OBJECTIVE: Although surgery in selected small-cell lung cancer (SCLC) patients has been proposed as a part of multimodality therapy, so far, the prognostic impact of node-spreading pattern has not been fully elucidated. To investigate this issue, a retrospective analysis was performed. METHODS: From 01/1996 to 12/2012, clinico-pathological, surgical, and oncological features were retrospectively reviewed in a multicentric cohort of 154 surgically treated SCLC patients. A multivariate Cox proportional hazard model was developed using stepwise regression, in order to identify independent outcome predictors. Overall (OS), cancer-specific (CSS), and Relapse-free survival (RFS) were calculated by Kaplan-Meier method. RESULTS: Overall, median OS, CSS, and RFS were 29 (95 % CI 18-39), 48 (95 % CI 19-78), and 22 (95 % CI 17-27) months, respectively. Lymphadenectomy was performed in 140 (90.9 %) patients (median number of harvested nodes: 11.5). Sixty-seven (47.9 %) pN0-cases experienced the best long-term survival (CSS: 71, RFS: 62 months; p < 0.0001). Among node-positive patients, no prognostic differences were found between pN1 and pN2 involvement (CSS: 22 vs. 15, and RFS: 14 vs. 10 months, respectively; p = 0.99). By splitting node-positive SCLC according to concurrent N1-invasion, N0N2-patients showed a worse CSS compared to those cases with combined N1N2-involvement (N0N2: 8 months vs. N1N2: 22 months; p = 0.04). On the other hand, the number of metastatic stations (p = 0.80) and the specific node-level (p = 0.85) did not affect CSS. At multivariate analysis, pN+ (HR: 3.05, 95 % CI 1.21-7.67, p = 0.02) and ratio between metastatic and resected lymph-nodes (RL, HR: 1.02, 95 % CI 1.00-1.04, p = 0.03) were independent predictors of CSS. Moreover, node-positive patients (HR: 3.60, 95 % CI 1.95-6.63, p < 0.0001) with tumor size ≥5 cm (HR: 1.85, 95 % CI 0.88-3.88, p = 0.10) experienced a worse RFS. CONCLUSIONS: In selected surgically treated SCLC, the long-term survival may be stratified according to the node-spreading pattern.

First-trimester euploid miscarriages analysed by array-CGH.

It is estimated that 10-15 % of all clinically recognised pregnancies results in a miscarriage, most of which occur during the first trimester. Large-scale chromosomal abnormalities have been found in up to 50 % of first-trimester spontaneous abortions and, for several decades, standard cytogenetic analysis has been used for their identification. Recent studies have proven that array comparative genomic hybridisation (array-CGH) is a useful tool for the detection of genome imbalances in miscarriages, showing a higher resolution, a significantly higher detection rate and overcoming problems of culture failures, maternal contamination and poor chromosome morphology. In this study, we investigated the possibility that submicroscopic chromosomal changes, not detectable by conventional cytogenetic analysis, exist in euploid miscarriages and could be causative for the spontaneous abortion. We analysed with array-CGH technology 40 foetal tissue samples derived by first-trimester miscarriages with a normal karyotype. A whole-genome microarray with a 100-Kb resolution was used for the analysis. Forty-five copy number variants (CNVs), ranging in size between 120 Kb and 4.3 Mb, were identified in 31 samples (24 gains and 21 losses). Ten samples (10/31, 32 %) have more than one CNV. Thirty-one CNVs (68 %) were defined as common CNVs and 14 were classified as unique. Six genes and five microRNAs contained within these CNVs will be discussed. This study shows that array-CGH is useful for detecting submicroscopic CNVs and identifying candidate genes which could account for euploid miscarriages.

Array-CGH and clinical characterization in a patient with subtelomeric 6p deletion without ocular dysgenesis.

Subtelomeric terminal 6p deletion has been recognized as a clinically identifiable syndrome including facial dysmorphism, malformation of the anterior eye chamber, hearing loss, heart defects, and developmental delay. Genotype-phenotype correlations of previously published patients have strongly suggested anterior eye segment anomalies as one of the major malformations of the syndrome if the critical 6p25 region contains the FOXC 1 gene. In addition, the presence in this region of one or more genes involved in hearing loss has been hypothesized. We report a patient with a 47,XYY karyotype and submicroscopic terminal 6p deletion. Further characterization of the deletion with array comparative genome hybridization also revealed a cryptic microduplication on chromosome 19. The patient showed dysmorphic features, neuromotor retardation, and profound language impairment, in absence of hearing loss and structural eye anomalies. As far as we know this is the first reported terminal 6p25.1 deletion case without eye dysgenesis precisely characterized by array-CGH. Our result suggests that the genes in this region may not be obvious candidates for hearing loss and demonstrate the need for further elucidation of the function of the genes involved in eye developmental processes.

The Italian external quality assessment scheme in classical cytogenetics: four years of activity.

BACKGROUND: The Italian external quality assessment scheme in classical cytogenetics was started in 2001 as an activity funded by the National Health System and coordinated by the Italian Public Institute of Health. OBJECTIVES: The aim of our work is to present data from the first 4 years of activity, 2001-2004. METHODS: Italian cytogenetics public laboratories were enrolled on a voluntary basis, and this nationwide program covered prenatal, postnatal and oncological diagnosis. The scheme is annual and retrospective; a panel of experts reviewed the quality of images and reports in order to assess technical, analytical and interpretative performance. RESULTS: Over the 4-year period, the number of participating laboratories increased: from 36 in 2001, 46 in 2002, 49 in 2003 to 51 in 2004. The overall technical performance was satisfactory. Inadequacy or lack of information in reporting was the most frequent analytical inaccuracy identified in all parts of the scheme. However, the percentage of complete reports increased significantly during the period: by 36% in postnatal diagnosis between 2001 and 2004 (p < 0.001) and by 42% in oncological diagnosis between 2002 and 2004 (p = 0.003). CONCLUSIONS: Our experience reveals that participation in external quality assessment programs has significant advantages, helping to standardize and to assure quality in cytogenetic testing.

Cryptic chromosome deletions involving SCN1A in severe myoclonic epilepsy of infancy.

OBJECTIVE: To identify cryptic chromosomal deletions involving SCN1A in patients with severe myoclonic epilepsy of infancy (SMEI). METHODS: Thirty-nine patients with SMEI and without SCN1A point mutations and their parents were typed with 14 intragenic SCN1A polymorphisms to identify hemizygosity. The parental origin and the extent of genomic deletions were determined by fluorescence in situ hybridization analysis using genomic clones encompassing chromosome 2q24.3-q31.1. Deletion breakpoints were more finely mapped by typing single-nucleotide polymorphisms and microsatellite markers. RESULTS: We identified three patients with SMEI who had genomic deletions encompassing the SCN1A locus. Deletion size was between 607 kb and 4.7 Mb. Deletions originated de novo from paternal chromosome in all subjects. One patient had central precocious puberty and palatoschisis. Genotype-phenotype correlations suggest that these clinical features are due to genes centromeric to SCN1A. CONCLUSIONS: Patients with severe myoclonic epilepsy of infancy (SMEI) lacking SCN1A point mutations should be investigated for cryptic chromosomal deletions involving SCN1A. Clinical features other than epilepsy could be associated with SMEI as a consequence of deletions in contiguous genes.

Mutation analysis of the NSD1 gene in a group of 59 patients with congenital overgrowth.

Sotos syndrome is characterized by pre- and post-natal overgrowth, typical craniofacial features, advanced bone age, and developmental delay. Some degree of phenotypic overlap exists with other overgrowth syndromes, in particular with Weaver syndrome. Sotos syndrome is caused by haploinsufficiency of the NSD1 (nuclear receptor SET domain containing gene 1) gene. Microdeletions involving the gene are the major cause of the syndrome in Japanese patients, whereas intragenic mutations are more frequent in non-Japanese patients. NSD1 aberrations have also been described in some patients diagnosed as Weaver syndrome. Some authors have suggested a certain degree of genotype-phenotype correlation, with a milder degree of overgrowth, a more severe mental retardation, and a higher frequency of congenital anomalies in microdeleted patients. Data on larger series are needed to confirm this suggestion. We report here on microdeletion and mutation analysis of NSD1 in 59 patients with congenital overgrowth. Fourteen novel mutations, two previously described and one microdeletion were identified. All patients with a NSD1 mutation had been clinically classified as "classical Sotos," although their phenotype analysis demonstrated that some major criteria, such as overgrowth and macrocephaly, could be absent. All patients with confirmed mutations shared the typical Sotos facial gestalt. A high frequency of congenital heart defects was present in patients with intragenic mutations, supporting the relevance of the NSD1 gene in the pathogenesis of this particular defect.

Cryptic 1p36.3/6q25.2 translocation in three generations ascertained through a foetus with IUGR and cerebral malformations.

Here we describe a foetus with intrauterine growth retardation (IUGR), cerebral malformations and a 46,XY,der(1),t(1;6)(p36.3;q25.2) karyotype owing to a familial cryptic translocation segregating in three generations. A balanced translocation was present in the mother, the maternal uncle, the aunt and the grandmother. A female first cousin with dysmorphisms, hydrocephalus and mental retardation was a carrier of a partial trisomy 1p and a partial monosomy 6q. Multiple miscarriages were present in the family pedigree. Parents of the foetus had three other pregnancies: a male with a balanced translocation, and two foetuses with 1p36.3-pter monosomy and 6q25.2-qter trisomy.

Review article: indications for anti-reflux surgery in gastro-oesophageal reflux disease.

Gastro-oesophageal reflux disease (GERD) is a complex multifactorial disorder whose treatment is based on knowledge of its pathophysiology, natural history and evolution. Recently the relationship between the severest degrees of cardial incontinence and hiatus hernia has been emphasized, which causes the impairment of the mechanical properties of the gastro-oesophageal barrier and of oesophageal acid clearing. Among different types of hiatus hernia, those characterized by the permanent axial orad migration of the oesophago-gastric (EG) junction (nonreducible hiatus hernia) are correlated with severe GERD. Barium swallow may adequately differentiate hiatal insufficiency, concentric hiatus hernia and short oesophagus which are the steps of migration across or above the diaphragm. When associated with panmural oesophagitis and fibrosis of the oesophageal wall, these conditions may be the cause of recurrence of hiatus hernia and reflux after laparoscopic standard anti-reflux surgical procedures; in the presence of nonreducibility of the EG junction below the diaphragm without tension, dedicated surgical procedures are necessary. It is currently agreed that surgical therapy is indicated for patients affected by severe GERD who are not compliant with long-term medical therapy, require high dosages of drugs and are too young for lifetime medical treatment. While the existence of severe GERD correlated with an irreversible anatomical disorder represents an elective indication for surgery, warrants further investigation. Accurate identification of the functional and anatomical abnormalities underlying GERD is mandatory in order to decide whether medical or surgical therapy should be implemented, and to tailor the surgical technique, laparoscopic or open, to each patient.

18q-syndrome and ectodermal dysplasia syndrome: description of a child and his family.

The 18q- syndrome [MIM #601808] is a terminal deletion of the long arm of chromosome 18. The most common deletion extends from region q21 to qter. We report here a nine-year-old boy possessing a simple 18q- deletion who had abnormalities of the brain, skull, face, tooth, hair, bone, and skin, plus joint laxity, tongue palsy, subtle sensoneural deafness, mental and speech delay, attention deficit hyperactivity disorder (ADHD), tic, and restless legs syndromes. His karyotype was 46, XY, del (18)(q21.31-qter). The size of the deletion was approximately 45 cM. Most of these abnormalities were not explained by the 18q- deletion. The family pedigree suggested the presence of a subtle involvement of ectodermal and/or mesodermal structures. Karyotypes of the other family members were normal.

Corpus callosum agenesis, multiple cysts, skin defects, and subtle ocular abnormalities with a de novo mutation [45,XX,der(5), t(5;;14) (pter;q11.2)].

We report on a 2-year-old girl with a de novo mutation [45,XX,der(5),t(5;14) (pter;q11.2)] with corpus callosum agenesis, multiple cysts (cerebral and cardiac), subtle eye abnormalities, and at least two different skin defects, strongly indicating neuroectodermal involvement, as a neuromuscular choristoma (hamartoma) and an eccrine hamartoma. Fluorescent in situ hybridization with different single-locus probes showed that chromosome 5 has a very small deletion, confined to a region composed of repetitive sequences. By contrast, the long (q) arm of chromosome 14 seems to be much more involved in the rearrangement, with partial monosomy spanning from the centromere to the D14S72 and D14S261 loci. The extent of the deleted region of chromosome 14 is approximately 16 cM. To our knowledge, this is the smallest reported deletion involving the chromosome 14q11.2 region to be associated with a developmental disorder resulting in variable eye, skin, and brain anomalies. We suggest that a new syndrome, mimicking in some ways the MLS phenotype, is caused by a deletion in the chromosome 14q11.2 region.

Clinical and molecular characterisation of 80 patients with 5p deletion: genotype-phenotype correlation.

The majority of deletions of the short arm of chromosome 5 are associated with cri du chat syndrome (CdCS) and patients show phenotypic and cytogenetic variability. To perform a genotype-phenotype correlation, 80 patients from the Italian CdCS Register were analysed. Molecular cytogenetic analysis showed that 62 patients (77.50%) had a 5p terminal deletion characterised by breakpoint intervals ranging from p13 (D5S763) to p15.2 (D5S18). Seven patients (8.75%) had a 5p interstitial deletion, four (5%) a de novo translocation, and three (3.75%) a familial translocation. Of the remaining four patients, three (3.75%) had de novo 5p anomalies involving two rearranged cell lines and one (1.25%) had a 5p deletion originating from a paternal inversion. The origin of the deleted chromosome 5 was paternal in 55 out of 61 patients (90.2%). Genotype-phenotype correlation in 62 patients with terminal deletions highlighted a progressive severity of clinical manifestation and psychomotor retardation related to the size of the deletion. The analysis of seven patients with interstitial deletions and one with a small terminal deletion confirmed the existence of two critical regions, one for dysmorphism and mental retardation in p15.2 and the other for the cat cry in p15.3. Results from one patient permitted the cat cry region to be distally narrowed from D5S13 to D5S731. Furthermore, this study lends support to the hypothesis of a separate region in p15.3 for the speech delay.

FISH characterization of two supernumerary r(1) associated with distinct clinical phenotypes.

Only a few reports on supernumerary r(1) chromosomes associated with a clinical phenotype have been published. We describe two unrelated patients with congenital malformations and developmental delay who were found to have a de novo supernumerary r(1) in 50% (Case 1) and 80% (Case 2) of the examined cells. Conventional cytogenetic techniques (QFQ, CBG, and DA-DAPI), complemented by fluorescence in situ hybridization studies using alpha satellite probes, showed that both small marker chromosomes (SMCs) primarily consisted of the centromere and heterochromatin of chromosome 1, a conclusion that was also supported by chromosome 1 painting. In an attempt to establish phenotype-genotype correlations, a further investigation was performed using YACs mapped to the chromosome 1 pericentromeric region. A fluorescent signal was evident after hybridization with Y934G9 (1q21) in Case 1 and Y959C4 (1p11.1-12) in Case 2. Partial trisomy of unique sequences flanking pericentromeric sequences is shown to underlie the clinical phenotype in both patients. This evidence should be taken into account when SMCs are ascertained, particularly in prenatal diagnosis.

Cytogenetic and molecular study of 32 Down syndrome families: potential leukaemia predisposing role of the most proximal segment of chromosome 21q.

Down syndrome (DS) children have a 10-20-fold increased risk of developing ALL or AML compared to non-DS children. An increased disomic homozygosity of the polymorphic DNA markers in the pericentromeric region of chromosome 21q (21q11) has repeatedly been found in DS patients with ANLL-M7 and DS-specific transient abnormal myelopoiesis (TAM), compared to the majority of DS subjects without leukaemia. Analysis of cytogenetic heteromorphisms and 26 polymorphic DNA markers from chromosome 21q showed an increased number of pericentromeric crossovers between the non-disjoined chromosomes in DS-ANLL cases (3/11), compared to DS-ALL (0/9) and DS-nonleukaemic cases (0/12). These findings are compatible with the model of disomic homozygosity of the predisposing allele of a putative pericentromeric gene, as an explanation for the high prevalence of ANLL in DS.

Double partial trisomy 9q34.1-->qter and 21pter-->q22.11: FISH and clinical findings.

We describe a patient with double trisomy 9q34.1-->qter and 21pter-->q22.1 resulting from 3:1 segregation of a maternal balanced translocation. The patient shows a clinical syndrome similar to that observed in patients with duplication of the chromosome 9q distal region, while no signs of trisomy 21 were observed. The use of high resolution banding and FISH were of fundamental importance for the cytogenetic diagnosis and for definition of the breakpoints on both chromosomes 9 and 21.

Combined use of cytogenetic analysis and FISH for the identification of two antenatal de novo markers as Robertsonian translocations involving the p arms.

In two prenatal cases, de novo nonmosaic bisatellited marker chromosomes were studied with the combined use of fluorescence in situ hybridization (FISH) with chromosome specific probes and cytogenetic heteromorphisms. The FISH studies showed that one of the small accessory chromosome could be a heterozygous 14/15 or 15/22 translocation involving the p arms of these chromosomes, the other showed only one hybridization spot with the classical satellite probe of chromosome 15. The analysis of heteromorphisms of the parental acrocentric chromosomes demonstrated that the two markers were Robertsonian translocations involving in the first case the p arms of the maternal 15 and 22 chromosomes and in the second case the p arms of the maternal 14 and 15 chromosomes.