Hutchinson Gilford Progeria Syndrome: A Therapeutic Approach via Adenoviral Delivery of CRISPR/cas Genome Editing System

Hutchinson-Gilford Progeria Syndrome (HGPS) is a rare human genetic disease caused by mutations in the LMNA gene. LMNA codes for structural components of the nuclear lamina. Alterations of nuclear lamina lead to a very variable class of diseases known as laminopathies. In detail, HGPS manifests a severe premature ageing phenotype due to the accumulation of a dominant negative form of lamin-A called progerin. With current treatments, the life expectancy of HGPS patients does not exceed their second decade. Death is usually due to cardiovascular complications. Recently, a new technology for mammals in vivo gene editing has been developed: the clustered regularly interspaced short palindromic repeats/Cas protein (CRISPR/Cas) system. The CRISPR/Cas technology permits to edit the genome at specific loci. Even if the CRSIPR/Cas constructs are transiently administered to the target cells, the genome editing is permanent. The advantages of the combination of non-integrating transient vectors in combination with the CRISPR/Cas constructs could give rise to a secure approach for the treatment of disease of genetic origin, especially those caused by dominant negative mutations, such as HGPS. A potential application of non-integrating transient vectors carrying CRISPR/Cas constructs for the treatment of HGPS will be discussed in detail

A ceRNA approach may unveil unexpected contributors to deletion syndromes, the model of 5q- syndrome

In genomic deletions, gene haploinsufficiency might directly configure a specific disease phenotype. Nevertheless, in some cases no functional association can be identified between haploinsufficient genes and the deletion-associated phenotype. Transcripts can act as microRNA sponges. The reduction of transcripts from the hemizygous region may increase the availability of specific microRNAs, which in turn may exert in-trans regulation of target genes outside the deleted region, eventually contributing to the phenotype. Here we prospect a competing endogenous RNA (ceRNA) approach for the identification of candidate genes target of epigenetic regulation in deletion syndromes. As a model, we analyzed the 5q- myelodysplastic syndrome. Genes in haploinsufficiency within the common 5q deleted region in CD34+ blasts were identified in silico. Using the miRWalk 2.0 platform, we predicted microRNAs whose availability, and thus activity, could be enhanced by the deletion, and performed a genomewide analysis of the genes outside the 5q deleted region that could be targeted by the predicted miRNAs. The analysis pointed to two genes with altered expression in 5q- transcriptome, which have never been related with 5q- before. The prospected approach allows investigating the global transcriptional effect of genomic deletions, possibly prompting discovery of unsuspected contributors in the deletion-associated phenotype. Moreover, it may help in functionally characterizing previously reported unexpected interactions

Anaplastic Thyroid Carcinoma: A ceRNA Analysis Pointed to a Crosstalk between SOX2

It has been suggested that cancer stem cells (CSC) may play a central role in oncogenesis, especially in undifferentiated tumours. Anaplastic thyroid carcinoma (ATC) has characteristics suggestive of a tumour enriched in CSC. Previous studies suggested that the stem cell factor SOX2 has a preeminent hierarchical role in determining the characteristics of stem cells in SW1736 ATC cell line. In detail, silencing SOX2 in SW1736 is able to suppress the expression of the stem markers analysed, strongly sensitizing the line to treatment with chemotherapeutic agents. Therefore, in order to further investigate the role of SOX2 in ATC, a competing endogenous RNA (ceRNA) analysis was conducted in order to isolate new functional partners of SOX2. Among the interactors, of particular interest are genes involved in the biogenesis of miRNAs (DICER1, RNASEN, and EIF2C2), in the control cell cycle (TP53, CCND1), and in mitochondrial activity (COX8A). The data suggest that stemness, microRNA biogenesis and functions, p53 regulatory network, cyclin D1, and cell cycle control, together with mitochondrial activity, might be coregulated

Competing Endogenous RNA: The Key to Posttranscriptional Regulation

Competing endogenous RNA, ceRNA, vie with messenger RNAs (mRNAs) for microRNAs (miRNAs) with shared miRNAs responses elements (MREs) and act as modulator of miRNA by influencing the available level of miRNA. It has recently been discovered that, apart from protein-coding ceRNAs, pseudogenes, long noncoding RNAs (lncRNAs), and circular RNAs act as miRNA “sponges” by sharing common MRE, inhibiting normal miRNA targeting activity on mRNA. These MRE sharing elements form the posttranscriptional ceRNA network to regulate mRNA expression. ceRNAs are widely implicated in many biological processes. Recent studies have identified ceRNAs associated with a number of diseases including cancer. This brief review focuses on the molecular mechanism of ceRNA as part of the complex post-transcriptional regulatory circuit in cell and the impact of ceRNAs in development and disease

Competing Endogenous RNAs, Non-Coding RNAs and Diseases: An Intertwined Story

MicroRNAs (miRNAs), a class of small non-coding RNA molecules, are responsible for RNA silencing and post-transcriptional regulation of gene expression. They can mediate a fine-tuned crosstalk among coding and non-coding RNA molecules sharing miRNA response elements (MREs). In a suitable environment, both coding and non-coding RNA molecules can be targeted by the same miRNAs and can indirectly regulate each other by competing for them. These RNAs, otherwise known as competing endogenous RNAs (ceRNAs), lead to an additional post-transcriptional regulatory layer, where non-coding RNAs can find new significance. The miRNA-mediated interplay among different types of RNA molecules has been observed in many different contexts. The analyses of ceRNA networks in cancer and other pathologies, as well as in other physiological conditions, provide new opportunities for interpreting omics data for the field of personalized medicine. The development of novel computational tools, providing putative predictions of ceRNA interactions, is a rapidly growing field of interest. In this review, I discuss and present the current knowledge of the ceRNA mechanism and its implications in a broad spectrum of different pathologies, such as cardiovascular or autoimmune diseases, cancers and neurodegenerative disorders

Are There Common Mechanisms Between the Hutchinson–Gilford Progeria Syndrome and Natural Aging?

The Hutchinson–Gilford progeria syndrome (HGPS) is a premature aging disease caused by mutations of the LMNA gene leading to increased production of a partially processed form of the nuclear fibrillar protein lamin A – progerin. Progerin acts as a dominant factor that leads to multiple morphological anomalies of cell nuclei and disturbances in heterochromatin organization, mitosis, DNA replication and repair, and gene transcription. Progerin-positive cells are present in primary fibroblast cultures obtained from the skin of normal donors at advanced ages. These cells display HGPS-like defects in nuclear morphology, decreased H3K9me3 and HP1, and increased histone H2AX phosphorylation marks of the DNA damage loci. Inhibition of progerin production in cells of aged non-HGPS donors in vivo increases the proliferative activity, H3K9me3, and HP1, and decreases the senescence markers p21, IGFBP3, and GADD45B to the levels of young donor cells. Thus, progerin-dependent mechanisms act in natural aging. Excessive activity of the same mechanisms may well be the cause of premature aging in HGPS. Telomere attrition is widely regarded to be one of the primary hallmarks of aging. Progerin expression in normal human fibroblasts accelerates the loss of telomeres. Changes in lamina organization may directly affect telomere attrition resulting in accelerated replicative senescence and progeroid phenotypes. The chronological aging in normal individuals and the premature aging in HGPS patients are mediated by similar changes in the activity of signaling pathways, including downregulation of DNA repair and chromatin organization, and upregulation of ERK, mTOR, GH-IGF1, MAPK, TGFβ, and mitochondrial dysfunction. Multiple epigenetic changes are common to premature aging in HGPS and natural aging. Recent studies showed that epigenetic systems could play an active role as drivers of both forms of aging. It may be suggested that these systems translate the effects of various internal and external factors into universal molecular hallmarks, largely common between natural and accelerated forms of aging. Drugs acting at both natural aging and HGPS are likely to exist. For example, vitamin D3 reduces the progerin production and alleviates most HGPS features, and also slows down epigenetic aging in overweight and obese non-HGPS individuals with suboptimal vitamin D status

Síndrome de Hutchinson-Gilford : abordagem terapêutica e genética

Dissertação para obtenção do grau de Mestre no Instituto Universitário Egas MonizUma das mais conhecidas síndromes Progeróides é a Progéria, também designada por Síndrome de Hutchinson – Gilford (HGPS). A Síndrome de Hutchinson – Gilford trata-se de uma patologia genética autossómica dominante resultante de uma mutação no gene LMNA, localizado no cromossoma 1, no locus 1q21.2-q21.2, caraterizada por originar um envelhecimento prematuro, sete vezes mais avançado que o envelhecimento fisiológico normal. A nível epidemiológico esta é uma doença rara, afetando um a cada quatro a oito milhões de recém-nascidos. Até ao momento da elaboração desta monografia não existe cura para esta síndrome, sendo que a terapêutica realizada nos portadores de progéria limita-se a atuar nos sintomas da mesma.One of the most known progeroid syndromes is Progeria, also known as Hutchinson-Gilford Syndrome (HGPS). Hutchinson-Gilford Syndrome is a dominant autosomal genetical disease resulting from a mutation in the LMNA gene, located on chromosome 1, locus 1q21.2-q21.2 characterized by a prematurely aging, about seven times faster than the normal physiological aging. At an epidemiological level this is a rare disease, affecting one in every four or eight newborns. By the time of elaboration of this monography there are no cure to this syndrome, and the therapy used in patients with progeria only treats the symptoms of the disease

Síndrome de Hutchinson-Gilford ou progéria: passado, presente e abordagens terapêuticas futuras

Dissertação de Mestrado, Ciências Farmacêuticas, Faculdade de Ciências e Tecnologia, Universidade do Algarve, 2014A síndrome de Hutchinson-Gilford (HGPS) ou progéria é uma doença rara e fatal que afeta 1 criança entre cada 4 milhões de nascimentos. Classificada como uma laminopatia progeróide segmental, induz alterações clínicas em múltiplos órgãos e tecidos, como estatura e peso corporal reduzidos, perda de cabelo precoce, lipodistrofia, escleroderma, osteólise e declínio cardiovascular que conduz a enfarte do miocárdio, a principal causa de morte nesta síndrome. No seu conjunto replicam fenótipos associados com o envelhecimento normal, sem afetar o seu desenvolvimentos cognitivo. Uma mutação autossómica dominante de novo GGC608GGT no gene LMNA causa a produção de uma prelamina A mutada, designada progerina. Por permanecer farnesilada, ancora-se ao invólucro nuclear. Além disso, devido à sua elevada expressão, acumula-se e altera a morfologia celular, a transição entre eucromatina e heterocromatinca fica comprometida e os telómeros disfuncionais, entre outras alterações, contribuem para a senescência celular e manifestação dos sintomas da doença. Após o diagnóstico confirmado pela expressão da maioria das caraterísticas clínicas e de um teste genético à mutação C→T específica da HGPS, as crianças e a respetiva família, acompanhadas pelo médico, poderão adotar medidas de suporte que permitem atenuar as consequências da doença. É comum a toma de Aspirina® de 2-3 mg/Kg de peso da criança, uma vez que a cura ainda não existe. Entretanto, a procura de uma terapia baseada na inibição do mecanismo pelo qual o grupo farnesilo é produzido e/ou se liga à progerina conduziu à realização de ensaios clínicos. Atualmente, está a ser testada uma combinação de lonafarnib, pravastatina e ácido zoledrónico, com conclusão prevista para 2017, mas já com relatos de melhoria em alguns aspetos do fenótipo HGPS. Novas abordagens terapêuticas serão necessárias para contemplar os restantes aspetos e descobrir-se a cura para estas crianças, que apesar de tudo são muito alegres e corajosas

A ceRNA analysis on LMNA gene focusing on the Hutchinson-Gilford progeria syndrome

Background: Hutchinson-Gilford progeria syndrome is a rare dominant human disease of genetic origin. The average life expectancy is about 20 years, patients’ life quality is still very poor and no efficient therapy has yet been developed. It is caused by mutation of the LMNA gene, which results in accumulation in the nuclear membrane of a particular splicing form of Lamin-A called progerin. The mechanism by which progerin perturbs cellular homeostasis and leads to the symptoms is still under debate. Micro-RNAs are able to negatively regulate transcription by coupling with the 3’ UnTranslated Region of messenger RNAs. Several Micro-RNAs recognize the same 3’ UnTranslated Region and each Micro-RNA can recognize multiple 3’ UnTranslated Regions of different messenger RNAs. When different messenger RNAs are co-regulated via a similar panel of micro-RNAs, these messengers are called Competing Endogenous RNAs, or ceRNAs. The 3’ UnTranslated Region of the longest LMNA transcript was analysed looking for its ceRNAs. The aim of this study was to search for candidate genes and gene ontology functions possibly influenced by LMNA mutations that may exert a role in progeria development. Results: 11 miRNAs were isolated as potential LMNA regulators. By computational analysis, the miRNAs pointed to 17 putative LMNA ceRNAs. Gene ontology analysis of isolated ceRNAs showed an enrichment in RNA interference and control of cell cycle functions. Conclusion: This study isolated novel genes and functions potentially involved in LMNA network of regulation that could be involved in laminopathies such as the Hutchinson-Gilford progeria syndrom