A Novel De novo Heterozygous Mutation in the SON Gene Associated with Septo-optic Dysplasia: A New Phenotype.

Septo-optic dysplasia (SOD) syndrome is a rare congenital disorder characterized by a classic triad of optic nerve/chiasm hypoplasia, agenesis of septum pellucidum and corpus callosum, and hypoplasia of the hypothalamic-pituitary axis.Herein, we report the clinical case of 2-year-old boy presenting with psychomotor delay, nystagmus, congenital hypothyroidism, and a clinically relevant growth delay. The neuroradiological examination showed partial segmental agenesis of the corpus callosum, agenesis of the septum pellucidum, optic nerve hypoplasia, and a small pituitary gland with a small median pituitary stalk. A whole-exome sequencing analysis detected a novel heterozygous de novo variant c.1069_1070delAG in SON, predicted as likely pathogenic.To date, SON pathogenic variants have been described as responsible for Zhu-Tokita-Takenouchi-Kim (ZTTK) syndrome, a multisystemic neurodevelopmental disorder mainly characterized by intellectual disability, facial dysmorphisms, visual abnormalities, brain malformations, feeding difficulties, and growth delay. The herein described case is the first recognized clinic-radiological occurrence of SOD syndrome with hypothalamic-pituitary dysfunction in a patient carrying a SON gene variant, considered responsible of ZTTK syndrome, suggesting a possible relationship between SOD and SON gene alterations, never described so far, making the search for SON gene mutations advisable in patients with SOD.

Overview of Curcumin and Piperine Effects on Glucose Metabolism: The Case of an Insulinoma Patient's Loss of Consciousness.

The hypoglycemic properties of curcumin supplements in therapeutic doses are well-known and may represent a useful tool for the treatment of chronic diseases such as metabolic syndrome, insulin resistance and type 2 diabetes. The poor bioavailability of curcumin can be improved with the concomitant administration of piperine, with no severe adverse effects on glycemia reported so far in the literature. In this article, we further discuss a previously reported case of a helicopter pilot, affected by grade I obesity who, under curcumin and piperine treatment, experienced a transient loss of consciousness (TLOC), during a low-altitude flight. This episode led to a diagnosis of insulinoma, previously asymptomatic. We hypothesized that the combined effects of curcumin and piperine might have caused a severe hypoglycemic episode and subsequent TLOC. Therefore, further studies should be conducted to evaluate the safety of curcumin and piperine supplementation in subjects with impaired glucose metabolism and insulin secretion.

Get Your Molar Tooth Right: Joubert Syndrome Misdiagnosis Unmasked by Whole-Exome Sequencing.

Joubert syndrome (JS) is a recessively inherited ciliopathy, characterized by a specific cerebellar and brainstem malformation recognizable on brain imaging as the "molar tooth sign" (MTS). Clinical signs include hypotonia, developmental delay, breathing abnormalities, and ocular motor apraxia. Older patients develop ataxia, intellectual impairment, and variable organ involvement. JS is genetically heterogeneous, with over 40 ciliary genes overall accounting for 65-75% cases. Thus, in recent years, the genetic diagnosis of JS has been based on the analysis of next-generation sequencing targeted gene panels. Since clinical features are unspecific and undistinguishable from other neurodevelopmental syndromes, the recognition of the MTS is crucial to address the patient to the appropriate genetic testing. However, the MTS is not always properly diagnosed, resulting either in false negative diagnoses (patients with the MTS not addressed to JS genetic testing) or in false positive diagnoses (patients with a different brain malformation wrongly addressed to JS genetic testing). Here, we present six cases referred for JS genetic testing based on inappropriate recognition of MTS. While the analysis of JS-related genes was negative, whole-exome sequencing (WES) disclosed pathogenic variants in other genes causative of distinct brain malformative conditions with partial clinical and neuroradiological overlap with JS. Reassessment of brain MRIs from five patients by a panel of expert pediatric neuroradiologists blinded to the genetic diagnosis excluded the MTS in all cases but one, which raised conflicting interpretations. This study highlights that the diagnostic yield of NGS-based targeted panels is strictly related to the accuracy of the diagnostic referral based on clinical and imaging assessment and that WES has an advantage over targeted panel analysis when the diagnostic suspicion is not straightforward.

Monoallelic KIF1A-related disorders: a multicenter cross sectional study and systematic literature review.

BACKGROUND: Monoallelic variants in the KIF1A gene are associated with a large set of clinical phenotypes including neurodevelopmental and neurodegenerative disorders, underpinned by a broad spectrum of central and peripheral nervous system involvement. METHODS: In a multicenter study conducted in patients presenting spastic gait or complex neurodevelopmental disorders, we analyzed the clinical, genetic and neuroradiological features of 28 index cases harboring heterozygous variants in KIF1A. We conducted a literature systematic review with the aim to comparing our findings with previously reported KIF1A-related phenotypes. RESULTS: Among 28 patients, we identified nine novel monoallelic variants, and one a copy number variation encompassing KIF1A. Mutations arose de novo in most patients and were prevalently located in the motor domain. Most patients presented features of a continuum ataxia-spasticity spectrum with only five cases showing a prevalently pure spastic phenotype and six presenting congenital ataxias. Seventeen mutations occurred in the motor domain of the Kinesin-1A protein, but location of mutation did not correlate with neurological and imaging presentations. When tested in 15 patients, muscle biopsy showed oxidative metabolism alterations (6 cases), impaired respiratory chain complexes II + III activity (3/6) and low CoQ10 levels (6/9). Ubiquinol supplementation (1gr/die) was used in 6 patients with subjective benefit. CONCLUSIONS: This study broadened our clinical, genetic, and neuroimaging knowledge of KIF1A-related disorders. Although highly heterogeneous, it seems that manifestations of ataxia-spasticity spectrum disorders seem to occur in most patients. Some patients also present secondary impairment of oxidative metabolism; in this subset, ubiquinol supplementation therapy might be appropriate.

Epilepsy in Tubulinopathy: Personal Series and Literature Review.

Mutations in tubulin genes are responsible for a large spectrum of brain malformations secondary to abnormal neuronal migration, organization, differentiation and axon guidance and maintenance. Motor impairment, intellectual disability and epilepsy are the main clinical symptoms. In the present study 15 patients from a personal cohort and 75 from 21 published studies carrying mutations in TUBA1A, TUBB2B and TUBB3 tubulin genes were evaluated with the aim to define a clinical and electrophysiological associated pattern. Epilepsy shows a wide range of severity without a specific pattern. Mutations in TUBA1A (60%) and TUBB2B (74%) and TUBB3 (25%) genes are associated with epilepsy. The accurate analysis of the Electroencephalogram (EEG) pattern in wakefulness and sleep in our series allows us to detect significant abnormalities of the background activity in 100% of patients. The involvement of white matter and of the inter-hemispheric connection structures typically observed in tubulinopathies is evidenced by the high percentage of asynchronisms in the organization of sleep activity recorded. In addition to asymmetries of the background activity, excess of slowing, low amplitude and Magnetic Resonance (MR) imaging confirm the presence of extensive brain malformations involving subcortical and midline structures. In conclusion, epilepsy in tubulinopathies when present has a favorable evolution over time suggesting a not particularly aggressive therapeutic approach.

Broadening the Spectrum of Adulthood X-Linked Adrenoleukodystrophy: A Report of Two Atypical Cases.

X-linked adrenoleukodystrophy (x-ALD) is a rare genetic disorder caused by a mutation in the ABCD1 gene, which encodes for a peroxisomal very long chain fatty acid transporter. Clinically, x-ALD can present a wide spectrum of different phenotypes: asymptomatic carriers, Addison only, cerebral x-ALD, and myelopathy with/without evidence of peripheral axonopathy (Adrenomyeloneuropathy). We report on two cases of adult x-ALD, with atypical phenotypes: (Case 1) A 37-years-old male with a 2-years-long history of spastic paraparesis, urinary urgency, and subclinical adrenocortical insufficiency. As an atypical finding, the MRI showed multiple congenital brain development defects. (Case 2) A 63-years-old male with a previous diagnosis of Addison disease, with a 6-years-long history of spastic paraparesis. Two years later, he complained of severe and disabling burning pain in his feet. A nerve conduction study was normal, but a skin biopsy revealed autonomic and somatic small fiber neuropathy. In both cases, genetic testing disclosed hemizygous mutation in ABCD1 associated with x-ALD: c.1394-2A > G and p.(Thr254Met), respectively. While case 1 supports the key role of peroxisome functions in brain development, case 2 points to a possible selective and clinically relevant peripheral small fiber degeneration in x-ALD myelopathy.

ZFYVE26/SPASTIZIN and SPG11/SPATACSIN mutations in hereditary spastic paraplegia types AR-SPG15 and AR-SPG11 have different effects on autophagy and endocytosis.

ZFYVE26/Spastizin and SPG11/Spatacsin encode 2 large proteins that are mutated in hereditary autosomal-recessive spastic paraplegia/paraparesis (HSP) type 15 (AR-SPG15) and type 11 (AR-SPG11), respectively. We previously have reported that AR-SPG15-related ZFYVE26 mutations lead to autophagy defects with accumulation of immature autophagosomes. ZFYVE26 and SPG11 were found to be part of a complex including the AP5 (adaptor related protein complex 5) and to have a critical role in autophagic lysosomal reformation with identification of autophagic and lysosomal defects in cells with both AR-SPG15- and AR-SPG11-related mutations. In spite of these similarities between the 2 proteins, here we report that ZFYVE26 and SPG11 are differently involved in autophagy and endocytosis. We found that both ZFYVE26 and SPG11 interact with RAB5A and RAB11, 2 proteins regulating endosome trafficking and maturation, but only ZFYVE26 mutations affected RAB protein interactions and activation. ZFYVE26 mutations lead to defects in the fusion between autophagosomes and endosomes, while SPG11 mutations do not affect this step and lead to a milder autophagy defect. We thus demonstrate that ZFYVE26 and SPG11 affect the same cellular physiological processes, albeit at different levels: both proteins have a role in autophagic lysosome reformation, but only ZFYVE26 acts at the intersection between endocytosis and autophagy, thus representing a key player in these 2 processes. Indeed expression of the constitutively active form of RAB5A in cells with AR-SPG15-related mutations partially rescues the autophagy defect. Finally the model we propose demonstrates that autophagy and the endolysosomal pathway are central processes in the pathogenesis of these complicated forms of hereditary spastic paraparesis. Abbreviations: ALR, autophagic lysosome reformation; AP5, adaptor related protein complex 5; AR, autosomal-recessive; HSP, hereditary spastic paraplegia/paraparesis; ATG14, autophagy related 14; BafA, bafilomycin A1; BECN1, beclin 1; EBSS, Earle balanced salt solution; EEA1, early endosome antigen 1; EGF, epidermal growth factor; EGFR, epidermal growth factor receptor; GDP, guanosine diphosphate; GFP, green fluorescent protein; GTP, guanosine triphosphate; HSP, hereditary spastic paraplegias; LBPA, lysobisphosphatidic acid; MAP1LC3B/LC3B, microtubule associated protein 1 light chain 3 beta; MVBs, multivesicular bodies; PIK3C3, phosphatidylinositol 3-kinase, catalytic subunit type 3; PIK3R4, phosphoinositide-3-kinase regulatory subunit 4; PtdIns3P, phosphatidylinositol-3-phosphate; RFP, red fluorescent protein; RUBCN, RUN and cysteine rich domain containing beclin 1 interacting protein; shRNA, short hairpin RNA; SQSTM1/p62, sequestosome 1; TCC: thin corpus callosum; TF, transferrin; UVRAG, UV radiation resistance associated.

Tubulin-related cerebellar dysplasia: definition of a distinct pattern of cerebellar malformation.

OBJECTIVE: To determine the neuroimaging pattern of cerebellar dysplasia (CD) and other posterior fossa morphological anomalies associated with mutations in tubulin genes and to perform clinical and genetic correlations. METHODS: Twenty-eight patients harbouring 23 heterozygous pathogenic variants (ten novel) in tubulin genes TUBA1A (n = 10), TUBB2B (n = 8) or TUBB3 (n = 5) were studied by a brain MRI scan performed either on a 1.5 T (n = 10) or 3 T (n = 18) MR scanner with focus on the posterior fossa. RESULTS: Cerebellar anomalies were detected in 24/28 patients (86%). CD was recognised in 19/28 (68%) including cortical cerebellar dysplasia (CCD) in 18/28, either involving only the cerebellar hemispheres (12/28) or associated with vermis dysplasia (6/28). CCD was located only in the right hemisphere in 13/18 (72%), including four TUBB2B-, four TUBB3- and five TUBA1A-mutated patients, while in the other five TUBA1A cases it was located only in the left hemisphere or in both hemispheres. The postero-superior region of the cerebellar hemispheres was most frequently affected. CONCLUSIONS: The cerebellar involvement in tubulinopathies shows specific features that may be labelled as 'tubulin-related CD'. This pattern is unique and differs from other genetic causes of cerebellar dysplasia. KEY POINTS: • Cortical cerebellar dysplasia without cysts is suggestive of tubulin-related disorder. • Cerebellar dysplasia in tubulinopathies shows specific features labelled as 'tubulin-related CD'. • Focal and unilateral involvement of cerebellar hemispheres has important implications for counselling.

Chromosomal imbalances in human bladder urothelial carcinoma: similarities and differences between biopsy samples and cancer stem-like cells.

BACKGROUND: The existence of two distinct groups of tumors with different clinical characteristic is a remarkable feature of transitional cell carcinomas (TCCs) of the bladder. More than 70% are low-grade (LG) non-infiltrating (NI) cancers at diagnosis, but 60-80% of them recur at least one time and 10-20% progress in stage and grade. On the other hand, about 20% of tumors show muscle invasion (IN) and have a poor prognosis with <50% survival after 5 years. This study focuses on the complexity of the bladder cancer genome, and for the first time to our knowledge, on the possibility to compare genomic alterations of in vitro selected cancer stem-like cells (CSCs), and their original biopsy in order to identify different genomic signature already present in the early stages of tumorigenesis of LG and HG tumors. METHODS: We initially used conventional chromosome analysis on TCC biopsies with different histotypes (LG vs HG) in order to detect rough differences between them. Then, we performed array comparative genomic hybridization (aCGH) on 10 HG and 10 LG tumors providing an overview of copy number alterations (CNAs). Finally, we made a comparison of the overall CNAs in 16 biopsies and their respective CSCs isolated from them. RESULTS: Our findings indicate that LG and HG bladder cancer differ with regard to their genomic profile even in the early stage of tumorigenesis; moreover, we identified a subgroup of LG samples with a higher tendency to lose genomic regions which could represent a more aggressive phenotype. CONCLUSIONS: The outcomes not only provide valuable information to deeper studying TCC carcinogenesis, but also could help in the clinic for diagnosis and prognosis of patients who will benefit from a more aggressive therapy.

DNA copy number alterations and PPARG amplification in a patient with multifocal bladder urothelial carcinoma.

BACKGROUND: Bladder cancer is the seventh most common cancer worldwide and over 90% are transitional cell carcinoma (TCC). At the first time of diagnosis at least 70% of TCC present as superficial bladder cancer. Because the clinical outcome of superficial bladder tumors is relatively unpredictable, there is a pressing need to identify markers that may predict tumor recurrence and progression and new treatment strategies. CASE PRESENTATION: We present a unique case of a 67-year old male who underwent total cystectomy after repeated trans-urethral resections of the bladder for multifocal non-muscle invasive bladder cancer. The first and the third tumor were diagnosed as high grade non-infiltrating (HGNI), while the second as carcinoma in situ (CIS). We performed both array comparative genomic hybridization and a targeted chromosomal profile by UroVysion in order to detect copy number variations (CNVs) that may be involved with tumor recurrence and progression. The overall data from this study provide new evidence for the monoclonal origin of urothelial tumor multifocality as several genetic changes were found in different tumors of the same patient. From the analysis of shared CNVs two gained regions emerged at 3p25.2 and 12q23.2, including PPARG and ASCL1 genes, respectively. The copy number level of these genes would seem inversely mutually correlated and highly dependent on histological grade, because the highest level of amplification at 3p25.2 was evidenced in the two HGNI samples, while the highest level of copy number gain at 12q23.2 was reported in the CIS. CONCLUSION: We provide new evidence on the role of PPARG in initiation and maintenance of bladder cancer. For the first time we also suggest a possible explanation for the elevated expression of PPARG in this type of tumor through a focal high level amplification at 3p25.2. Furthermore, a new gene, ASCL1, emerged as a potential candidate to assist PPARG in bladder carcinogenesis.

Chromosomal aberrations in bladder cancer: fresh versus formalin fixed paraffin embedded tissue and targeted FISH versus wide microarray-based CGH analysis.

Bladder carcinogenesis is believed to follow two alternative pathways driven by the loss of chromosome 9 and the gain of chromosome 7, albeit other nonrandom copy number alterations (CNAs) were identified. However, confirmation studies are needed since many aspects of this model remain unclear and considerable heterogeneity among cases has emerged. One of the purposes of this study was to evaluate the performance of a targeted test (UroVysion assay) widely used for the detection of Transitional Cell Carcinoma (TCC) of the bladder, in two different types of material derived from the same tumor. We compared the results of UroVysion test performed on Freshly Isolated interphasic Nuclei (FIN) and on Formalin Fixed Paraffin Embedded (FFPE) tissues from 22 TCCs and we didn't find substantial differences. A second goal was to assess the concordance between array-CGH profiles and the targeted chromosomal profiles of UroVysion assay on an additional set of 10 TCCs, in order to evaluate whether UroVysion is an adequately sensitive method for the identification of selected aneuploidies and nonrandom CNAs in TCCs. Our results confirmed the importance of global genomic screening methods, that is array based CGH, to comprehensively determine the genomic profiles of large series of TCCs tumors. However, this technique has yet some limitations, such as not being able to detect low level mosaicism, or not detecting any change in the number of copies for a kind of compensatory effect due to the presence of high cellular heterogeneity. Thus, it is still advisable to use complementary techniques such as array-CGH and FISH, as the former is able to detect alterations at the genome level not excluding any chromosome, but the latter is able to maintain the individual data at the level of single cells, even if it focuses on few genomic regions.

Biological heterogeneity of putative bladder cancer stem-like cell populations from human bladder transitional cell carcinoma samples.

Transitional cell carcinoma (TCC) is the most common type of bladder cancer. Emerging evidence has suggested that the capability of a tumor to grow and propagate is dependent on a small subset of cells, the cancer stem-like cells (CSCs) or tumor initiating cells. We report on the isolation and biological characterization of putative bladder CSC populations from primary TCCs. Isolated cells were induced to proliferate in stem cell culture conditions (serum-free medium containing mitogenic growth factors). The proliferating cells formed spheroids (urospheres) and their abilities for extensive proliferation and self-renewal were assayed. Their positivity for several stem cell markers (CD133, Oct-3/4, nestin, and cytokeratins) was also assessed by immunofluorescence tests and they could have the potential to differentiate in the presence of serum. In stem cell culture conditions they gradually showed loss of proliferation, adherence to the substrate, and morphological changes, which might reflect their progressive acquisition of differentiative capacity and loss of self-renewal ability. To evaluate if effective cell selection occurred after isolation, conventional cytogenetic studies on fresh chromosome spreads immediately after isolation and after culture were carried out. In addition, a molecular cytogenetic study by UroVysion assay was carried out on paraffin-embedded tissue sections and on fresh and after culture nuclei preparations. The data collected indicated important karyotype changes and a positive selection for hypo- or near-diploid cells, losing the complexity present in fresh tumors.