Identificazione e analisi funzionale delle mutazioni rilevate nel gene ANKRD26 responsabili di una nuova forma di piastrinopenia ereditaria

2011/2012La Trombocitopenia 2, anche definita come ANKRD26-Related Disease (ANKRD26-RD), rientra fra le nuove forme di piastrinopenie ereditarie recentemente caratterizzate. Si tratta di una trombocitopenia non sindromica a trasmissione autosomica dominante causata da mutazione nel gene ANKRD26. Fra le diverse caratteristiche cliniche spicca un’aumentata incidenza di sviluppare leucemie fra i pazienti affetti. Data l’associazione con il cancro, si rileva la necessità da una parte di migliorare la diagnosi delle diverse forme per comprenderne il meccanismo patogenetico e dall’altra di caratterizzare il fenotipo clinico della malattia indagando sul ruolo delle mutazioni a carico di ANKRD26 nell’alterare la produzione di piastrine e nello sviluppo di neoplasie. Abbiamo, pertanto, analizzato ANKRD26 in una casistica di pazienti piastrinopenici privi di una diagnosi identificando diverse mutazioni in eterozigosi, tutte localizzate nella regione 5’ non tradotta del gene. I pazienti affetti presentano una piastrinopenia moderata; i sintomi più comuni sono petecchie, ecchimosi, epistassi e menorragia. È stata evidenziata una diminuzione degli alfa granuli e un aumento della trombopoietina sierica. Nel midollo è stata osservata un’importante dismegacariocitopoiesi. E’ emerso infine, dagli studi fin qui condotti, una correlazione tra ANKRD26-RD e la leucemia: infatti l’incidenza di sviluppare leucemia acuta, nei pazienti con ANKRD26-RD, è di 167 casi su 100000 un’incidenza più alta che nel resto della popolazione stimata fra i 3,4 e 6,6 casi su 100000 (National Cancer institute). Per quanto riguarda la ricerca volta a definire i meccanismi patogenetici della malattia, abbiamo ipotizzato che le mutazioni, vista la loro localizzazione in una regione regolatrice, alterassero i meccanismi di controllo dell’espressione di ANKRD26 durante la megacariocitopoiesi. Come primo passo, abbiamo eseguito un’analisi bioinformatica per ricercare eventuali fattori di trascrizioni che riconoscono la regione delle mutazioni. L’analisi ha evidenziato un sito di legame per il fattore RUNX1. Successivamente abbiamo eseguito dei saggi di luciferasi attraverso i quali abbiamo dimostrato come le mutazioni determinino un aumento statisticamente significativo dell’attività di luciferasi almeno in cellule megacarioblastiche, DAMI, che possono essere differenziate in megacariociti. Infine abbiamo confermato mediante saggi EMSA l’interazione della regione delle mutazioni con il fattore RUNX1 predetto dal programma bioinformatico. Per due mutazioni (c.-128G>A e c.-119C>A), inoltre, abbiamo individuato un altro fattore di trascrizione HMGA1a non predetto dall’analisi bioinformatica. Infine studi di espressione su CD34+ e CD41+ hanno mostrato che ANKRD26 è maggiormente espresso nelle cellule staminali CD34+ ed è meno espresso nei megacariociti CD41+, lasciandoci ipotizzare che durante il processo di differenziamento ci sia una riduzione dei livelli di espressione del gene. Tutti questi dati ci suggeriscono, quindi, che il 5’UTR sia coinvolto nelle regolazione di ANKRD26. Saranno avviate, pertanto, nuove indagini volte ad caratterizzare ulteriormente i meccanismi mediante i quali le mutazioni interferiscono con il controllo dell’espressione, permettendoci di comprendere l’eziopatogenesi e la caratterizzazione della ANKRD26-RD.XXV Cicl

Microduplication in the 2p16.1p15 chromosomal region linked to developmental delay and intellectual disability

Abstract Background Several patients with the 2p16.1p15 microdeletion syndrome have been reported. However, microduplication in the 2p16.1p15 chromosomal region has only been reported in one case, and milder clinical features were present compared to those attributed to 2p16.1p15 microdeletion syndrome. Some additional cases were deposited in DECIPHER database. Case presentation In this report we describe four further cases of 2p16.1p15 microduplication in four unrelated probands. They presented with mild gross motor delay, delayed speech and language development, and mild dysmorphic features. In addition, two probands have macrocephaly and one a congenital heart anomaly. Newly described cases share several phenotype characteristics with those detailed in one previously reported microduplication case. Conclusion The common features among patients are developmental delay, speech delay, mild to moderate intellectual disability and unspecific dysmorphic features. Two patients have bilateral clinodactyly of the 5th finger and two have bilateral 2nd-3rd toes syndactyly. Interestingly, as opposed to the deletion phenotype with some cases of microcephaly, 2 patients are reported with macrocephaly. The reported cases suggest that microduplication in 2p16.1p15 chromosomal region might be causally linked to developmental delay, speech delay, and mild intellectual disability

International collaboration as a tool for diagnosis of patients with inherited thrombocytopenia in the setting of a developing country

Inherited thrombocytopenias (ITs) are heterogeneous genetic disorders that frequently represent a diagnostic challenge. The requirement of highly specialized tests for diagnosis represents a particular problem in resourcelimited settings. To overcome this difficulty, we applied a diagnostic algorithm and developed a collaboration program with a specialized international center in order to increase the diagnostic yield in a cohort of patients in Argentina. Methods: Based on the algorithm, initial evaluation included collection of clinical data, platelet size, blood smear examination and platelet aggregation tests. Confirmatory tests were performed according to diagnostic suspicion, which included platelet glycoprotein expression, immunofluorescence for myosin- 9 in granulocytes and platelet thrombospondin-1 and molecular screening of candidate genes. Results: Thirty-one patients from 14 pedigrees were included; their median age was 32 (4?72) years and platelet count 72 (4?147) · 109 L)1. Autosomal dominant inheritance was found in nine (64%) pedigrees; 10 (71%) had large platelets and nine (29%) patients presented with syndromic forms. A definitive diagnosis was made in 10 of 14 pedigrees and comprised MYH9-related disease in four, while classic and monoallelic Bernard?Soulier syndrome, gray platelet syndrome, X-linked thrombocytopenia, thrombocytopenia 2 (ANKRD26 mutation) and familial platelet disorder with predisposition to acute myelogenous leukemia were diagnosed in one pedigree each. Conclusions: Adoption of an established diagnostic algorithm and collaboration with an expert referral center proved useful for diagnosis of IT patients in the setting of a developing country. This initiative may serve as a model to develop international networks with the goal of improving diagnosis and care of patients with these rare diseases.Fil: Glembotsky, Ana Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Marta, Rosana Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Pecci, Alessandro. Universita Degli Studi Di Pavia; ItaliaFil: de Rocco, Daniela. Institute for Maternal and Child Health – IRCCS "Burlo Garofolo"; ItaliaFil: Gnan, Chiara. Institute for Maternal and Child Health – IRCCS "Burlo Garofolo"; ItaliaFil: Espasandin, Yesica Romina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Goette, Nora Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Negro, F.. Instituto Médico Sagrado Corazón; ArgentinaFil: Noris, Patrizia. Universita Degli Studi Di Pavia; ItaliaFil: Savoia, Anna. Institute for Maternal and Child Health – IRCCS "Burlo Garofolo"; Italia. Università degli Studi di Trieste; ItaliaFil: Balduini, C. L.. Universita Degli Studi Di Pavia; ItaliaFil: Molinas, Felisa Concepción. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Heller, Paula Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; Argentin

Germline mutations in ETV6 are associated with thrombocytopenia, red cell macrocytosis and predisposition to lymphoblastic leukemia

Some familial platelet disorders are associated with predisposition to leukemia, myelodysplastic syndrome (MDS) or dyserythropoietic anemia. We identified a family with autosomal dominant thrombocytopenia, high erythrocyte mean corpuscular volume (MCV) and two occurrences of B cell-precursor acute lymphoblastic leukemia (ALL). Whole-exome sequencing identified a heterozygous single-nucleotide change in ETV6 (ets variant 6), c.641C>T, encoding a p.Pro214Leu substitution in the central domain, segregating with thrombocytopenia and elevated MCV. A screen of 23 families with similar phenotypes identified 2 with ETV6 mutations. One family also had a mutation encoding p.Pro214Leu and one individual with ALL. The other family had a c.1252A>G transition producing a p.Arg418Gly substitution in the DNA-binding domain, with alternative splicing and exon skipping. Functional characterization of these mutations showed aberrant cellular localization of mutant and endogenous ETV6, decreased transcriptional repression and altered megakaryocyte maturation. Our findings underscore a key role for ETV6 in platelet formation and leukemia predisposition

"The why and how of managerialization of family businesses: evidences from Italy"

This paper concerns how management processes adopted by family firms are influenced by family’s and organizational characteristics; more specifically we would like to contribute to the debate on the managerialization of family firms, by studying the determinants of the adoption of managerial systems. We consider managerialization of family business related to the diffusion of formal managerial mechanisms, both strategic planning and managerial control systems, and human resource management systems. We distinguish between determinants of managerialization related the family’s characteristics, such as involvement of family/non family managers, and organizational drivers. Differently from previous studies, we articulated family involvement in management, considering separately the involvement of family members in the top management team, in the techno-structure, and in the middle management. The research hypotheses have been tested on a sample of 99 family firms from the Novara province, in Italy. Findings highlight that more organizational and strategic complexity, than family involvement in management explain the diffusion of managerial systems. They also show that the presence of a family CFO and a family HR manager (techno-structure) is positively associated with the adoption of formal managerial mechanisms. Our results suggest that in any firm (both family and not family one, of any size, operating both in manufacturing and not manufacturing industries) when firm’s complexity increases both managerial roles, especially in terms of techno-structure, and managerial mechanisms have to be introduced

Role of the mutations identified in the 5\u2019UTR of ANKRD26 responsible for an inherited form of thrombocytopenia

8nononenoneC. Gnan; L. Arnoldo; D.De Rocco; M. Cusan; R. Sgarra; M. Faleschini; G. Manfioletti; A. SavoiaGnan, Chiara; Arnoldo, Laura; DE ROCCO, Daniela; Cusan, Martina; Sgarra, Riccardo; Faleschini, Michela; Manfioletti, Guidalberto; Savoia, Ann