Abstract Background/Rationale. Hereditary Angioedema (HAE) is a rare genetic disease (estimated prevalence of 1:50,000) that causes recurrent local edema with important negative impact on the patiens’ quality of life and risk of death when the larynx is affected. Angioedema is mediated by bradykinin, released when a triggering factor, mainly physical and psychological stress, activates the contact system. Treatment is only partially satisfactory. Knowledge and awarness for HAE are very limited. The disease is caused by functional C1-inhibitor (C1−INH) deficiency in plasma, which is due to mutations in one of the two alleles coding for this protein and it is the biochemical marker of the disease. Plasma levels of functional C1−INH are constantly markedly below the half normal value that the normal allele should provide and that would prevent angioedema symptoms. Angioedema pathogenesis is only partially defined and triggering mechanisms totally unknown. A down regulation of C1−INH normal allele can be relevant to the development of angioedema (PAPPAL). Description of the project. The project is aimed to provide a better understanding on the pathogenesis of symptoms and on the regulation of C1−INH gene in order to provide targets for new therapeutic approaches to cure this neglected, severe genetic disease. Specifically, angioedema pathogenesis will be approached with genomic mRNA expression studies by microaray to define the involved genes and plasma protein studies to detect those biochemical changes that mark an angioedema attack. C1−INH expression will be analyzed by mRNA quantitation in patients’ cells and by translational and post−translational analysis of C1−INH gene regulation. Specific aims. a. to identify the genes involved in angioedema attacks; b. to identify the biochemical changes that occur in plasma and mark an angioedema attack; c. to investigate if a down regulation of normal C1−INH allele causes functional C1−INH levels below 50% of normal and exposes HAE patients to the risk of angioedema attacks. Anticipated output. The project can define new genes involved in the pathogenesis of angioedema and a down regulation of wild type C1−INH allele as the condition that expose HAE patients to develop angioedema. Proximity to cure. The project is focused on the identification of new approaches to the therapy of HAE. The treatments available at present are focused on directly antagonizing or preventing the release of the mediator of symptoms (CIC). With our approach we will investigate 2 new possibilities that are closer to the complete cure of the disease. One is to identify genes other that C1−INH that intervene in the triggering of angioedema. The other evaluates the existence and the mechanism causing a functional repression of the normal allele. Thus our project will not immediately result in new therapies, but in the identification of new targets to be exploited for new therapies. Role and contribution of partners in the project. The project is the result of the cooperation of 3 centers A (Marco Cicardi, Milan), B (Giuseppe Castellano, Bari), C (Sonia Caccia, Milano). Center A has important specific background on HAE. The world’s largest case list of these patients is followed at this center that will have the role of coordination providing all patients’ samples. Center A will also take the responsibility of the studies with tilt test stimulation that will be designed and performed by physicians with specific experience for studies on the regulation of the sympathetic nervous system. The same center will perform the studies on patients’ plasma and preparation of RNA samples. Cenetr B will have the responsibility of the microarray studies. The group has some specific experience on HAE patients since an outpatient clinic for HAE has been active for 6 years in their Department. The background on microarray analysis is based on previous publications on other disease such as chronic kidney disease and leukemia. Center C will be in charge of all the studies of gene expression regulation both in terms of real−time RT−PCR quantitation of allele specific C1−INH mRNA levels in patients’ samples and in terms of in−vitro studies of transcriptional and post−transcriptional regulation of gene expression. Center C has a consolidated experience in studying the molecular basis of inherited human genetic diseases, specifically, in: i) the analysis of gene−expression regulation at the transcriptional and post−transcriptional level, ii) the study of the effect of mutations at the m−RNA level and iii) the functional and structural characterization of mutant proteins expressed in heterologous expression−systems. Project summary for general public (in Italian) L’angioedema ereditario (HAE) è una malattia genetica rara causa di edemi ricorrenti localizzati alla cute, alla mucosa del tratto gastrointestinale e alla laringe. Queste manifestazioni hanno un importante impatto negativo sulla qualità di vita dei pazienti affetti e nel caso in cui vi sia un coinvolgimento della laringe possono portare a morte per asfissia. Il mediatore degli attacchi di angioedema è la bradichinina che viene rilasciata quando fattori scatenanti, quali per esempio stress fisici e psicologici, attivano il sistema di contatto. I trattamenti disponibili sono solo parzialmente soddisfacenti. La carenza di C1−inibitore (C1−INH), che è alla base della malattia, è dovuta a mutazioni in uno dei due alleli che codificano per questa proteina. I livelli funzionali di C1−INH nel plasma dei pazienti con HAE sono sempre inferiori alla metà del valore normale. L’allele sano dovrebbe tuttavia provvedere a mantenere dei valori di C1−INH intorno alla metà dei valori normali e quindi prevenire i sintomi di angioedema. La patogenesi del HAE è, quindi, solo parzialmente definita e i meccanismi che scatenano gli attacchi ancora completamente sconosciuti. Una “down regulation” dell’allele sano per C1−INH potrebbe essere rilevante nello sviluppo dell’angioedema. Questo progetto è mirato ad identificare nuovi geni coinvolti nella patogenesi del HAE e nella “down regulation” dell’allele sano del C1−INH per chiarire la condizione che espone i pazienti affetti da HAE a sviluppare attacchi di angioedema. Questi risultati possono essere di grande rilevanza nella sviluppo di nuove terapie. In particolare, la patogenesi dell’angioedema sarà indagata attraverso lo studio delle proteine plasmatiche per indagare i cambiamenti biochimici durante un attacco di angioedema, con studi di espressione dell’mRNA attraverso metodiche di microarray per definire i geni coinvolti e con studi di espressione genica nelle cellule dei pazienti ed in opportuni sistemi cellulari per approfondirne il meccanismo molecolare.

Gene regulation in the pathogenesis of angioedema due to inherited c1 inhibitor deficiency (hereditary angioedema) / M. Cicardi, A. Zanichelli, E. Bonanni, C. Suffritti, L. Maggioni, S. Caccia, R. Russo, Y. Zhi, S. Duga, R. Asselta, V. Rimoldi, G. Castellano. ((Intervento presentato al 15. convegno Telethon Scientific Convention tenutosi a Riva del Garda nel 2009.

Gene regulation in the pathogenesis of angioedema due to inherited c1 inhibitor deficiency (hereditary angioedema)

M. Cicardi;A. Zanichelli;C. Suffritti;L. Maggioni;S. Caccia;R. Russo;ZHI, YUXIANG;S. Duga;R. Asselta;V. Rimoldi;G. Castellano
2009-03-07

Abstract

Abstract Background/Rationale. Hereditary Angioedema (HAE) is a rare genetic disease (estimated prevalence of 1:50,000) that causes recurrent local edema with important negative impact on the patiens’ quality of life and risk of death when the larynx is affected. Angioedema is mediated by bradykinin, released when a triggering factor, mainly physical and psychological stress, activates the contact system. Treatment is only partially satisfactory. Knowledge and awarness for HAE are very limited. The disease is caused by functional C1-inhibitor (C1−INH) deficiency in plasma, which is due to mutations in one of the two alleles coding for this protein and it is the biochemical marker of the disease. Plasma levels of functional C1−INH are constantly markedly below the half normal value that the normal allele should provide and that would prevent angioedema symptoms. Angioedema pathogenesis is only partially defined and triggering mechanisms totally unknown. A down regulation of C1−INH normal allele can be relevant to the development of angioedema (PAPPAL). Description of the project. The project is aimed to provide a better understanding on the pathogenesis of symptoms and on the regulation of C1−INH gene in order to provide targets for new therapeutic approaches to cure this neglected, severe genetic disease. Specifically, angioedema pathogenesis will be approached with genomic mRNA expression studies by microaray to define the involved genes and plasma protein studies to detect those biochemical changes that mark an angioedema attack. C1−INH expression will be analyzed by mRNA quantitation in patients’ cells and by translational and post−translational analysis of C1−INH gene regulation. Specific aims. a. to identify the genes involved in angioedema attacks; b. to identify the biochemical changes that occur in plasma and mark an angioedema attack; c. to investigate if a down regulation of normal C1−INH allele causes functional C1−INH levels below 50% of normal and exposes HAE patients to the risk of angioedema attacks. Anticipated output. The project can define new genes involved in the pathogenesis of angioedema and a down regulation of wild type C1−INH allele as the condition that expose HAE patients to develop angioedema. Proximity to cure. The project is focused on the identification of new approaches to the therapy of HAE. The treatments available at present are focused on directly antagonizing or preventing the release of the mediator of symptoms (CIC). With our approach we will investigate 2 new possibilities that are closer to the complete cure of the disease. One is to identify genes other that C1−INH that intervene in the triggering of angioedema. The other evaluates the existence and the mechanism causing a functional repression of the normal allele. Thus our project will not immediately result in new therapies, but in the identification of new targets to be exploited for new therapies. Role and contribution of partners in the project. The project is the result of the cooperation of 3 centers A (Marco Cicardi, Milan), B (Giuseppe Castellano, Bari), C (Sonia Caccia, Milano). Center A has important specific background on HAE. The world’s largest case list of these patients is followed at this center that will have the role of coordination providing all patients’ samples. Center A will also take the responsibility of the studies with tilt test stimulation that will be designed and performed by physicians with specific experience for studies on the regulation of the sympathetic nervous system. The same center will perform the studies on patients’ plasma and preparation of RNA samples. Cenetr B will have the responsibility of the microarray studies. The group has some specific experience on HAE patients since an outpatient clinic for HAE has been active for 6 years in their Department. The background on microarray analysis is based on previous publications on other disease such as chronic kidney disease and leukemia. Center C will be in charge of all the studies of gene expression regulation both in terms of real−time RT−PCR quantitation of allele specific C1−INH mRNA levels in patients’ samples and in terms of in−vitro studies of transcriptional and post−transcriptional regulation of gene expression. Center C has a consolidated experience in studying the molecular basis of inherited human genetic diseases, specifically, in: i) the analysis of gene−expression regulation at the transcriptional and post−transcriptional level, ii) the study of the effect of mutations at the m−RNA level and iii) the functional and structural characterization of mutant proteins expressed in heterologous expression−systems. Project summary for general public (in Italian) L’angioedema ereditario (HAE) è una malattia genetica rara causa di edemi ricorrenti localizzati alla cute, alla mucosa del tratto gastrointestinale e alla laringe. Queste manifestazioni hanno un importante impatto negativo sulla qualità di vita dei pazienti affetti e nel caso in cui vi sia un coinvolgimento della laringe possono portare a morte per asfissia. Il mediatore degli attacchi di angioedema è la bradichinina che viene rilasciata quando fattori scatenanti, quali per esempio stress fisici e psicologici, attivano il sistema di contatto. I trattamenti disponibili sono solo parzialmente soddisfacenti. La carenza di C1−inibitore (C1−INH), che è alla base della malattia, è dovuta a mutazioni in uno dei due alleli che codificano per questa proteina. I livelli funzionali di C1−INH nel plasma dei pazienti con HAE sono sempre inferiori alla metà del valore normale. L’allele sano dovrebbe tuttavia provvedere a mantenere dei valori di C1−INH intorno alla metà dei valori normali e quindi prevenire i sintomi di angioedema. La patogenesi del HAE è, quindi, solo parzialmente definita e i meccanismi che scatenano gli attacchi ancora completamente sconosciuti. Una “down regulation” dell’allele sano per C1−INH potrebbe essere rilevante nello sviluppo dell’angioedema. Questo progetto è mirato ad identificare nuovi geni coinvolti nella patogenesi del HAE e nella “down regulation” dell’allele sano del C1−INH per chiarire la condizione che espone i pazienti affetti da HAE a sviluppare attacchi di angioedema. Questi risultati possono essere di grande rilevanza nella sviluppo di nuove terapie. In particolare, la patogenesi dell’angioedema sarà indagata attraverso lo studio delle proteine plasmatiche per indagare i cambiamenti biochimici durante un attacco di angioedema, con studi di espressione dell’mRNA attraverso metodiche di microarray per definire i geni coinvolti e con studi di espressione genica nelle cellule dei pazienti ed in opportuni sistemi cellulari per approfondirne il meccanismo molecolare.
C1-inhibitor ; Hereditary Angioedema
Settore BIO/11 - Biologia Molecolare
Settore MED/09 - Medicina Interna
Telethon
Gene regulation in the pathogenesis of angioedema due to inherited c1 inhibitor deficiency (hereditary angioedema) / M. Cicardi, A. Zanichelli, E. Bonanni, C. Suffritti, L. Maggioni, S. Caccia, R. Russo, Y. Zhi, S. Duga, R. Asselta, V. Rimoldi, G. Castellano. ((Intervento presentato al 15. convegno Telethon Scientific Convention tenutosi a Riva del Garda nel 2009.
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