Diana Postorivo

Four Copies of SNCA Responsible for Autosomal Dominant Parkinson's Disease in Two Italian Siblings.

Background. Parkinsons disease (PD) is mostly characterized by alpha-synuclein (SNCA) aggregation and loss of nigrostriatal dopamine-containing neurons. In this study a novel SNCA multiplication is described in two siblings affected by severe parkinsonism featuring early onset dyskinesia, psychiatric symptoms, and cognitive deterioration. Methods. SNCA dosage was performed using High-Density Comparative Genomic Hybridization Array (CGH-Array), Multiple Ligation Dependent Probe Amplification (MLPA), and Quantitative PCR (qPCR). Genetic analysis was associated with clinical evaluation. Results. Genetic analysis of siblings showed for the first time a 351 Kb triplication containing SNCA gene along with 6 exons of MMRN1 gene in 4q22.1 and a duplication of 1,29 Mb of a genomic region flanking the triplication. Conclusions. The identification of this family indicates a novel mechanism of SNCA gene multiplication, which confirms the genomic instability in this region and provides data on the genotype-phenotype correlation in PD patients.

Oct-1 recruitment to the nuclear envelope in adult-onset autosomal dominant leukodystrophy

Adult-onset autosomal dominant leukodystrophy (ADLD) is a slowly progressive neurological disorder characterised by pyramidal, cerebellar, and autonomic disturbances. Duplication of the LMNB1 gene is the genetic cause of ADLD, yet the pathogenetic mechanism is not defined. In this study, we analysed cells and muscle tissue from three patients affected by ADLD, carrying an extra copy of the LMNB1 gene. Lamin B1 levels were dramatically increased in ADLD nuclei, both in skin fibroblasts and skeletal muscle fibres. Since lamin B1 is known to bind Oct-1, a transcription factor involved in the oxidative stress pathway, we investigated Oct-1 fate in ADLD. Oct-1 recruitment to the nuclear periphery was increased in ADLD cells, while nucleoplasmic localisation of the transcription factor under oxidative stress conditions was reduced. Importantly, lamin B1 degradation occurring in some, but not all ADLD cell lines, slowed down lamin B1 and Oct-1 accumulation. In skeletal muscle, focal disorganisation of sarcomeres was observed, while IIB-myosin heavy chain, an Oct-1 target gene, was under-expressed and rod-containing fibres were formed. These data show that a high degree of regulation of lamin B1 expression is implicated in the different clinical phenotypes observed in ADLD and show that altered Oct-1 nuclear localisation contributes to the disease phenotype.

First evidence of Smith–Magenis syndrome in mother and daughter due to a novel RAI mutation

Smith–Magenis syndrome (SMS) is a complex genetic disorder caused by interstitial 17p11.2 deletions encompassing multiple genes, including the retinoic acid induced 1 gene—RAI1—or mutations in RAI1 itself. The clinical spectrum includes developmental delay, cognitive impairment, and behavioral abnormalities, with distinctive physical features that become more evident with age. No patients have been reported to have had offspring. We here describe a girl with developmental delay, mainly compromising the speech area, and her mother with mild intellectual disabilities and minor dysmorphic features. Both had sleep disturbance and attention deficit disorder, but no other atypical behaviors have been reported. In both, CGH‐array analysis detected a 15q13.3 interstitial duplication, encompassing CHRNA7. However, the same duplication has been observed in several, apparently healthy, maternal relatives. We, thus, performed a whole exome sequencing analysis, which detected a frameshift mutation in RAI1, de novo in the mother, and transmitted to her daughter. No other family members carried this mutation. This is the first report of an SMS patient having offspring. Our experience confirms the importance of searching for alternative causative genetic mechanisms in case of confounding/inconclusive findings such as a CGH‐array result of uncertain significance.

Easychip 8x15k: A New Tool for Detecting Chromosome Anomalies in Low Risk Pregnancies, Supporting and Integrating Standard Karyotype

Over last decade chromosome microarray analysis has become a routine test, but its use as first tier in prenatal diagnosis still raises disputes specially when applied to low risk pregnancies. In order to limit the identification of incidental findings (IF) and variants of unknown significance (VOUS) we designed EasyChip, a low-resolution oligonucleotide array CGH platform with a functional resolution of 3 Mb in genomic backbone, 300 Kb in sub-telomeric regions, and 150 Kb in 43 regions associated with syndromic disorders, selected considering morbidity, penetrance, and etiological mechanisms. After an “in silico” evaluation, which showed that Easychip would not uncover most of VOUS (24% vs 3%) and any IF, we have validated EasyChip on 169 patients samples, 57 retrospective samples with known imbalances and 112 prospective samples as part of the prenatal diagnosis process. All the known rearrangements were detected and 7 further pathogenic imbalances were detected on the still undiagnosed cohort. To evaluate false positive/negative rate, thirty-eight out of the 112 prospective samples were also processed on an high resolution array CGH, allowing comparing the results in term of diagnostic utility and impact on detection rate. Two positive and pathogenic results were detected by both platforms. EasyChip did not detect 10 of the 11 VOUS nor 2 IF discovered by the high-resolution platform. In conjunction with karyotype, EasyChip is a useful tool in prenatal diagnosis for screening purposes on low risk pregnancies, it enables the detection of cryptic imbalanced subtelomeric rearrangements, microdeletions/duplications within 43 syndromic regions and supports standard cytogenetic analysis at whole genome level. Finally, this tool, differently from higher resolution platforms, significantly reduces the detection rate of VOUS and IF, which represent a major drawback during genetic counselling specially for low risk pregnancies, significantly reduces the time to spend on analysis and limit the need of additional confirmation.

A Small Supernumerary Marker Derived from the Pericentromeric Region of Chromosome 5: Case Report and Delineation of Partial Trisomy 5p Phenotype

A 17-year-old girl presented with a distinct phenotype mainly featuring craniofacial dysmorphism, including a disproportioned large, round, elongated face; hypertelorism; deep-set eyes with short palpebral fissures; obesity (BMI 37), and a neuropsychiatric disorder with high-functioning autism. Postnatal conventional cytogenetic analyses from peripheral blood revealed a mosaic small supernumerary marker chromosome (sSMC) with a mos 47,XX,+mar[7]/46,XX[43] karyotype. By cenM-FISH technique, the sSMC was identified as a ring derivative of chromosome 5. Metaphase FISH analysis with a set of dedicated probes defined its origin from the pericentromeric region of chromosome 5, including the NIPBL gene at 5p13.2. Such sSMCs, exceedingly rare in the literature, underlie proximal trisomy 5p. In order to delineate a core phenotype of proximal trisomy 5p, we compared our patients features with those of 6 patients found in the literature with similar der(5) chromosomes. Furthermore, a dozen individuals with 5p13 (micro)duplication syndrome was compared and discussed. We identified highly distinctive craniofacial dysmorphism, obesity, and intellectual disability and/or autism spectrum disorder as typical features of proximal 5p trisomy. In the critical region (band 5p13), the NIPBL gene is likely to be a major determinant of the neurobehavioral phenotype, and its presence at the sSMC level may be relevant to predict clinical outcome.

Transabdominal coelocentesis as early source of fetal DNA for chromosomal and molecular diagnosis

Abstract This study reports a comparative analysis between results of transabdominal coelocentesis and traditional invasive procedure in order to assess the usefulness of coelocentesis as a source of fetal DNA for molecular and chromosomal analysis. A number of 28 women were included in the study. A successful sampling of coelomic fluid was obtained in 25 women by transabdominal procedure. A positive amplification of DNA with QF-PCR techniques was obtained in 90% of cases, while 10% of cases failed to reveal interpretable results. Although all samples were cultured, the growth rate was not sufficient to determine karyotypes within 2 weeks. Five samples were selected to be analyzed by array-based comparative genomic hybridization (a-CGH) but the interpretation of these results was difficult and ambiguous. Our results suggest that transabdominal coelocentesis is suitable for the detection of single DNA variation and for QF-PCR analysis, while further experiments are needed to develop optimized protocols for traditional karyotyping and array-analysis.

Copy number variations in healthy subjects. Case study: iPSC line CSSi005-A (3544) production from an individual with variation in 15q13.3 chromosome duplicating gene CHRNA7

CHRNA7, encoding the neuronal alpha7 nicotinic acetylcholine receptor (a7nAChR), is highly expressed in the brain, particularly in the hippocampus. It is situated in the 15q13.3 chromosome region, frequently associated with a Copy Number Variation (CNV), which causes its duplication or deletion. The clinical significance of CHRNA7 duplications is unknown so far, but there are several research data suggesting that they may be pathogenic, with reduced penetrance. We have produced an iPS cell line from a single healthy donors fibroblasts carrying a 15q13.3 CNV, including CHRNA7 in order to study the exact role of this CNV during the neurodevelopment.

Small 4p16.3 deletions: Three additional patients and review of the literature

Wolf–Hirschhorn syndrome is a well‐defined disorder due to 4p16.3 deletion, characterized by distinct facial features, intellectual disability, prenatal and postnatal growth retardation, and seizures. Genotype–phenotype correlations based on differently sized deletions have been attempted, and some candidate genes have been suggested. We report on clinical characteristics of three patients with pure interstitial submicroscopic 4p16.3 deletions, ranging in size from 68 to 166 kb, involving WHSCR1 and/or part of WHSCR2, and review published cases with overlapping 4p16.3 losses. The present study highlights a major role of NSD2 gene in the pathogenesis of the WHS main features and predicts that loss‐of‐function mutations affecting NSD2 gene could result in microcephaly, prenatal and postnatal growth retardation, psychomotor and language delay, and craniofacial features. Absent seizures in all subjects corroborate the suggestion that this specific feature is causally linked with at least one additional causative gene. Finally, we suggest that mir‐943 could play a role in the pathogenesis of CHD in some of these patients.

Identification of i(X)(p10) as the sole molecular abnormality in atypical chronic myeloid leukemia evolved into acute myeloid leukemia

The World Health Organization classifies atypical chronic myeloid leukemia (aCML) as a myeloproliferative/myelodisplastic hematological disorder. The primary manifestations are leukocytosis with disgranulopoiesis, absence of basophilia and/or monocytosis, splenomegaly and absence of Philadelphia chromosome or BCR/ABL fusion. Overall 50-65% of patients demonstrate karyotypic abnormalities, although no specific cytogenetic alterations have been associated with this disease. X chromosome alterations have been rarely reported in myeloid malignancies. Although Isodicentric X, idic(X)(q13) is well known in females with myelodysplastic syndromes (MDS), little data are available on X isochromosome and its pathogenetic potential in these disorders. i(X)(p10) is observed in a variety of hematologic malignancies, both myeloid and lymphoid, as a unique abnormality, as well as part of a more complex karyotype, in females and less frequently in male patients. The present report describes the first patient with aCML, with documented isolated i(X)(p10), who developed a secondary acute myeloid leukemia (sAML).

Molecular cytogenetics characterization of seven small supernumerary marker chromosomes derived from chromosome 19: Genotype-phenotype correlation and review of the literature

Only a few subjects carrying supernumerary marker chromosomes derived from 19 chromosome (sSMC(19)) have been described to date and for a small portion of them the genic content has been defined at the molecular level. We present seven new different sSMCs(19) identified in eight individuals, seven of whom unrelated. The presence of the sSMC is associated with a clinical phenotype in five subjects, while the other three carriers, two of whom related, are normal. All sSMCs(19) have been characterized by means of conventional and molecular cytogenetics. We compare the sSMCs(19) carriers with a clinical phenotype to already described patients with gains (sSMCs or microduplications) of overlapping genomic regions with the aim to deepen the pathogenicity of the encountered imbalances and to assess the role of the involved genes on the phenotype. The present work supports the correlation between the gain of some chromosome 19 critical regions and specific phenotypes.