Tumor Suppressor Role of Wild-Type P53-Dependent Secretome and Its Proteomic Identification in PDAC

: The study of the cancer secretome is gaining even more importance in cancers such as pancreatic ductal adenocarcinoma (PDAC), whose lack of recognizable symptoms and early detection assays make this type of cancer highly lethal. The wild-type p53 protein, frequently mutated in PDAC, prevents tumorigenesis by regulating a plethora of signaling pathways. The importance of the p53 tumor suppressive activity is not only primarily involved within cells to limit tumor cell proliferation but also in the extracellular space. Thus, loss of p53 has a profound impact on the secretome composition of cancer cells and marks the transition to invasiveness. Here, we demonstrate the tumor suppressive role of wild-type p53 on cancer cell secretome, showing the anti-proliferative, apoptotic and chemosensitivity effects of wild-type p53 driven conditioned medium. By using high-resolution SWATH-MS technology, we characterized the secretomes of p53-deficient and p53-expressing PDAC cells. We found a great number of secreted proteins that have known roles in cancer-related processes, 30 of which showed enhanced and 17 reduced secretion in response to p53 silencing. These results are important to advance our understanding on the link between wt-p53 and cancer microenvironment. In conclusion, this approach may detect a secreted signature specifically driven by wild-type p53 in PDAC

Genetic and phenotypic spectrum associated with IFIH1 gain-of-function

IFIH1 gain-of-function has been reported as a cause of a type I interferonopathy encompassing a spectrum of autoinflammatory phenotypes including Aicardi–Goutières syndrome and Singleton Merten syndrome. Ascertaining patients through a European and North American collaboration, we set out to describe the molecular, clinical and interferon status of a cohort of individuals with pathogenic heterozygous mutations in IFIH1. We identified 74 individuals from 51 families segregating a total of 27 likely pathogenic mutations in IFIH1. Ten adult individuals, 13.5% of all mutation carriers, were clinically asymptomatic (with seven of these aged over 50 years). All mutations were associated with enhanced type I interferon signaling, including six variants (22%) which were predicted as benign according to multiple in silico pathogenicity programs. The identified mutations cluster close to the ATP binding region of the protein. These data confirm variable expression and nonpenetrance as important characteristics of the IFIH1 genotype, a consistent association with enhanced type I interferon signaling, and a common mutational mechanism involving increased RNA binding affinity or decreased efficiency of ATP hydrolysis and filament disassembly rate

The Need for Biomarkers in the ALS-FTD Spectrum: A Clinical Point of View on the Role of Proteomics

: Frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are severely debilitating and progressive neurodegenerative disorders. A distinctive pathological feature of several neurodegenerative diseases, including ALS and FTD, is the deposition of aberrant protein inclusions in neuronal cells, which leads to cellular dysfunction and neuronal damage and loss. Despite this, to date, the biological process behind developing these protein inclusions must be better clarified, making the development of disease-modifying treatment impossible until this is done. Proteomics is a powerful tool to characterize the expression, structure, functions, interactions, and modifications of proteins of tissue and biological fluid, including plasma, serum, and cerebrospinal fluid. This protein-profiling characterization aims to identify disease-specific protein alteration or specific pathology-based mechanisms which may be used as markers of these conditions. Our narrative review aims to highlight the need for biomarkers and the potential use of proteomics in clinical practice for ALS-FTD spectrum disorders, considering the emerging rationale in proteomics for new drug development. Certainly, new data will emerge in the near future in this regard and support clinicians in the development of personalized medicine

Tumor Suppressor Role of Wild-Type P53-Dependent Secretome and Its Proteomic Identification in PDAC

The study of the cancer secretome is gaining even more importance in cancers such as pancreatic ductal adenocarcinoma (PDAC), whose lack of recognizable symptoms and early detection assays make this type of cancer highly lethal. The wild-type p53 protein, frequently mutated in PDAC, prevents tumorigenesis by regulating a plethora of signaling pathways. The importance of the p53 tumor suppressive activity is not only primarily involved within cells to limit tumor cell proliferation but also in the extracellular space. Thus, loss of p53 has a profound impact on the secretome composition of cancer cells and marks the transition to invasiveness. Here, we demonstrate the tumor suppressive role of wild-type p53 on cancer cell secretome, showing the anti-proliferative, apoptotic and chemosensitivity effects of wild-type p53 driven conditioned medium. By using high-resolution SWATH-MS technology, we characterized the secretomes of p53-deficient and p53-expressing PDAC cells. We found a great number of secreted proteins that have known roles in cancer-related processes, 30 of which showed enhanced and 17 reduced secretion in response to p53 silencing. These results are important to advance our understanding on the link between wt-p53 and cancer microenvironment. In conclusion, this approach may detect a secreted signature specifically driven by wild-type p53 in PDAC

Astroglial calcium signaling and homeostasis in tuberous sclerosis complex

: Tuberous Sclerosis Complex (TSC) is a multisystem genetic disorder characterized by the development of benign tumors in various organs, including the brain, and is often accompanied by epilepsy, neurodevelopmental comorbidities including intellectual disability and autism. A key hallmark of TSC is the hyperactivation of the mechanistic target of rapamycin (mTOR) signaling pathway, which induces alterations in cortical development and metabolic processes in astrocytes, among other cellular functions. These changes could modulate seizure susceptibility, contributing to the progression of epilepsy and its associated comorbidities. Epilepsy is characterized by dysregulation of calcium (Ca2+) channels and intracellular Ca2+ dynamics. These factors contribute to hyperexcitability, disrupted synaptogenesis, and altered synchronization of neuronal networks, all of which contribute to seizure activity. This study investigates the intricate interplay between altered Ca2+ dynamics, mTOR pathway dysregulation, and cellular metabolism in astrocytes. The transcriptional profile of TSC patients revealed significant alterations in pathways associated with cellular respiration, ER and mitochondria, and Ca2+ regulation. TSC astrocytes exhibited lack of responsiveness to various stimuli, compromised oxygen consumption rate and reserve respiratory capacity underscoring their reduced capacity to react to environmental changes or cellular stress. Furthermore, our study revealed significant reduction of store operated calcium entry (SOCE) along with strong decrease of basal mitochondrial Ca2+ concentration and Ca2+ influx in TSC astrocytes. In addition, we observed alteration in mitochondrial membrane potential, characterized by increased depolarization in TSC astrocytes. Lastly, we provide initial evidence of structural abnormalities in mitochondria within TSC patient-derived astrocytes, suggesting a potential link between disrupted Ca2+ signaling and mitochondrial dysfunction. Our findings underscore the complexity of the relationship between Ca2+ signaling, mitochondria dynamics, apoptosis, and mTOR hyperactivation. Further exploration is required to shed light on the pathophysiology of TSC and on TSC associated neuropsychiatric disorders offering further potential avenues for therapeutic development

Encephalopathies with intracranial calcification in children: Clinical and genetic characterization

none42noBackground: We present a group of patients affected by a paediatric onset genetic encephalopathy with cerebral calcification of unknown aetiology studied with Next Generation Sequencing (NGS) genetic analyses. Methods: We collected all clinical and radiological data. DNA samples were tested by means of a customized gene panel including fifty-nine genes associated with known genetic diseases with cerebral calcification. Results: We collected a series of fifty patients. All patients displayed complex and heterogeneous phenotypes mostly including developmental delay and pyramidal signs and less frequently movement disorder and epilepsy. Signs of cerebellar and peripheral nervous system involvement were occasionally present. The most frequent MRI abnormality, beside calcification, was the presence of white matter alterations; calcification was localized in basal ganglia and cerebral white matter in the majority of cases. Sixteen out of fifty patients tested positive for mutations in one of the fifty-nine genes analyzed. In fourteen cases the analyses led to a definite genetic diagnosis while results were controversial in the remaining two. Conclusions: Genetic encephalopathies with cerebral calcification are usually associated to complex phenotypes. In our series, a molecular diagnosis was achieved in 32% of cases, suggesting that the molecular bases of a large number of disorders are still to be elucidated. Our results confirm that cerebral calcification is a good criterion to collect homogeneous groups of patients to be studied by exome or whole genome sequencing; only a very close collaboration between clinicians, neuroradiologists and geneticists can provide better results from these new generation molecular techniques.noneTonduti D.; Panteghini C.; Pichiecchio A.; Decio A.; Carecchio M.; Reale C.; Moroni I.; Nardocci N.; Campistol J.; Garcia-Cazorla A.; Perez Duenas B.; Zorzi G.; Ardissone A.; Granata T.; Freri E.; Zibordi F.; Ragona F.; D'Arrigo S.; Saletti V.; Esposito S.; Pantaleoni C.; Riva D.; De Giorgis V.; Cereda C.; Valente M.L.; Sproviero D.; Poo Arguelles M.P.; Estupina C.F.; Sans Fito A.M.; Martorell Sampol L.; Del Mar O'Callaghan Gordo M.; Ortez Gonzalez C.I.; Gonzalez Alvarez V.; Garcia-Segarra N.; Fusco C.; Bertini E.; Diodato D.; Fazzi E.; Galli J.; Chiapparini L.; Garavaglia B.; Orcesi S.Tonduti, D.; Panteghini, C.; Pichiecchio, A.; Decio, A.; Carecchio, M.; Reale, C.; Moroni, I.; Nardocci, N.; Campistol, J.; Garcia-Cazorla, A.; Perez Duenas, B.; Zorzi, G.; Ardissone, A.; Granata, T.; Freri, E.; Zibordi, F.; Ragona, F.; D'Arrigo, S.; Saletti, V.; Esposito, S.; Pantaleoni, C.; Riva, D.; De Giorgis, V.; Cereda, C.; Valente, M. L.; Sproviero, D.; Poo Arguelles, M. P.; Estupina, C. F.; Sans Fito, A. M.; Martorell Sampol, L.; Del Mar O'Callaghan Gordo, M.; Ortez Gonzalez, C. I.; Gonzalez Alvarez, V.; Garcia-Segarra, N.; Fusco, C.; Bertini, E.; Diodato, D.; Fazzi, E.; Galli, J.; Chiapparini, L.; Garavaglia, B.; Orcesi, S

Molecular Genetics and Interferon Signature in the Italian Aicardi Goutières Syndrome Cohort: Report of 12 New Cases and Literature Review

Aicardi-Goutières syndrome (AGS) is a genetically determined early onset encephalopathy characterized by cerebral calcification, leukodystrophy, and increased expression of interferon-stimulated genes (ISGs). Up to now, seven genes (TREX1, RNASEH2B, RNASEH2C, RNASEH2A, ADAR1, SAMHD1, IFIH1) have been associated with an AGS phenotype. Next Generation Sequencing (NGS) analysis was performed on 51 AGS patients and interferon signature (IS) was investigated in 18 AGS patients and 31 healthy controls. NGS identified mutations in 48 of 51 subjects, with three patients demonstrating a typical AGS phenotype but not carrying mutations in known AGS-related genes. Five mutations, in RNASEH2B, SAMHD1 and IFIH1 gene, were not previously reported. Eleven patients were positive and seven negatives for the upregulation of interferon signaling (IS > 2.216). This work presents, for the first time, the genetic data of an Italian cohort of AGS patients, with a higher percentage of mutations in RNASEH2B and a lower frequency of mutations in TREX1 than those seen in international series. RNASEH2B mutated patients showed a prevalence of negative IS consistent with data reported in the literature. We also identified five novel pathogenic mutations that warrant further functional investigation. Exome/genome sequencing will be performed in future studies in patients without a mutation in AGS-related genes

Molecular Genetics and Interferon Signature in the Italian Aicardi Gouti\ue8res Syndrome Cohort: Report of 12 New Cases and Literature Review

Aicardi-Goutieres syndrome (AGS) is a genetically determined early onset encephalopathy characterized by cerebral calcification, leukodystrophy, and increased expression of interferon-stimulated genes (ISGs). Up to now, seven genes (TREX1, RNASEH2B, RNASEH2C, RNASEH2A, ADAR1, SAMHD1, IFIH1) have been associated with an AGS phenotype. Next Generation Sequencing (NGS) analysis was performed on 51 AGS patients and interferon signature (IS) was investigated in 18 AGS patients and 31 healthy controls. NGS identified mutations in 48 of 51 subjects, with three patients demonstrating a typical AGS phenotype but not carrying mutations in known AGS-related genes. Five mutations, in RNASEH2B, SAMHD1 and IFIH1 gene, were not previously reported. Eleven patients were positive and seven negatives for the upregulation of interferon signaling (IS > 2.216). This work presents, for the first time, the genetic data of an Italian cohort of AGS patients, with a higher percentage of mutations in RNASEH2B and a lower frequency of mutations in TREX1 than those seen in international series. RNASEH2B mutated patients showed a prevalence of negative IS consistent with data reported in the literature. We also identified five novel pathogenic mutations that warrant further functional investigation. Exome/genome sequencing will be performed in future studies in patients without a mutation in AGS-related genes

Genetic and phenotypic spectrum associated with IFIH1 gain-of-function

IFIH1 gain-of-function has been reported as a cause of a type I interferonopathy encompassing a spectrum of autoinflammatory phenotypes including Aicardi-Goutières syndrome and Singleton Merten syndrome. Ascertaining patients through a European and North American collaboration, we set out to describe the molecular, clinical and interferon status of a cohort of individuals with pathogenic heterozygous mutations in IFIH1. We identified 74 individuals from 51 families segregating a total of 27 likely pathogenic mutations in IFIH1. Ten adult individuals, 13.5% of all mutation carriers, were clinically asymptomatic (with seven of these aged over 50 years). All mutations were associated with enhanced type I interferon signaling, including six variants (22%) which were predicted as benign according to multiple in silico pathogenicity programs. The identified mutations cluster close to the ATP binding region of the protein. These data confirm variable expression and nonpenetrance as important characteristics of the IFIH1 genotype, a consistent association with enhanced type I interferon signaling, and a common mutational mechanism involving increased RNA binding affinity or decreased efficiency of ATP hydrolysis and filament disassembly rate