Mostra na ABL comemora os 70 anos de Serney

Exposição vai mostrar ao público vida do escritor e político.

Caracterização do envolvimento do gene RECKna proliferação celular e progressão tumoral: inversa correlação com a expressão do oncogene c-myc

Este trabalho mostra o envolvimento do gene RECK no processo de progressão do ciclo celular. Foi verificado que a expressão endógena de RECK é modulada durante a progressão do ciclo celular. A superexpressão de RECK em fibroblastos normais de camundongo promove uma diminuição da capacidade proliferativa das células e um retardo da transição das fases G0/G1-S do ciclo celular. Além disso, os resultados sugerem que um dos possíveis mecanismos de ação de RECK, que promovem este processo, envolve a indução da expressão de um inibidor de CDK, especificamente de p21, e retardo da fosforilação de pRb. Os resultados indicam, ainda, que durante a progressão do ciclo celular a expressão do gene RECK apresenta uma correlação inversa com a expressão do proto-oncogene c-myc. Estes dados corroboram os dados da literatura que mostram RECK como um alvo para o produto de diversos oncogenes, como ras e c-myc. A caracterização da repressão de RECK por c-Myc mostrou que a mesma ocorre ao nível transcricional e que sítios Sp1, presentes no promotor de RECK, são essenciais para a ação de Myc. Dados adicionais sugerem que a repressão de RECK por c-Myc parece envolver mecanismos de desacetilação de histonas. A modulação da expressão de RECK também foi avaliada durante a progressão maligna de tumores do sistema nervoso central (especificamente, gliomas). Foi verificado que a expressão de RECK não é alterada com a progressão deste tipo de tumor. Porém, foi verificado que os pacientes que manifestaram um maior tempo de sobrevida apresentaram tumores com uma significativa maior expressão do gene RECK. Estes dados sugerem que RECK possa ser um possível marcador prognóstico. A caracterização da regulação da expressão de RECK, tanto em células normais como em diferentes tipos de tumores, assim como os alvos moleculares da sua ação, são pontos muito importantes para o entendimento dos mecanismos que controlam a proliferação celular e podem contribuir para o desenvolvimento de novas formas de terapia anti-tumoral.

Ḥabl al-matīn: rūznāmah-ʼi muqaddas

Description based on: Sāl-i 33., shumārah-i 3 (2 Farvarī sanah-i 1925 Mīlādī); title from masthead.

Selection pressure-driven evolution of the Epstein-Barr virus-encoded oncogene LMP1 in virus isolates from southeast Asia

The geographically constrained distribution of Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) in southeast Asian populations suggests that both viral and host genetics may influence disease risk. Although susceptibility loci have been mapped within the human genome, the role of viral genetics in the focal distribution of NPC remains an enigma. Here we report a molecular phylogenetic analysis of an NPC-associated viral oncogene, LMP1, in a large panel of EBV isolates from southeast Asia and from Papua New Guinea, Africa, and Australia, regions of the world where NPC is and is not endemic, respectively. This analysis revealed that LMP1 sequences show a distinct geographic structure, indicating that the southeast Asian isolates have evolved as a lineage distinct from those of Papua New Guinea, African, and Australian isolates. Furthermore, a likelihood ratio test revealed that the C termini of the LMP1 sequences of the southeast Asian lineage are under significant positive selection pressure, particularly at some sites within the C-terminal activator regions. We also present evidence that although the N terminus and transmembrane region of LMP1 have undergone recombination, the C-terminal region of the gene has evolved without any history of recombination. Based on these observations, we speculate that selection pressure may be driving the LMP1 sequences in virus isolates from southeast Asia towards a more malignant phenotype, thereby influencing the endemic distribution of NPC in this region.

Induction of Therapeutic T-Cell Responses to Subdominant Tumor-associated Viral Oncogene after Immunization with Replication-incompetent Polyepitope Adenovirus Vaccine

The EBV-encoded latent membrane proteins (LMP1 and LMP2), which are expressed in various EBV-associated malignancies have been proposed as a potential target for CTL-based therapy. However, the precursor frequency for LMP-specific CTL is generally low, and immunotherapy based on these antigens is often compromised by the poor immunogenicity and potential threat from their oncogenic potential. Here we have developed a replication-incompetent adenoviral vaccine that encodes multiple HLA class I-restricted CTL epitopes from LMP1 and LMP2 as a polyepitope. Immunization with this polyepitope vaccine consistently generated strong LMP-specific CTL responses in HLA A2/K-b mice, which can be readily detected by both ex vivo and in vivo T-cell assays. Furthermore, a human CTL response to LMP antigens can be rapidly expanded after stimulation with this recombinant polyepitope vector. These expanded T cells displayed strong lysis of autologous target cells sensitized with LMP1 and/or LMP2 CTL epitopes. More importantly, this adenoviral vaccine was also successfully used to reverse the outgrowth of LMP1-expressing tumors in HLA A2/K-b mice. These studies demonstrate that a replication-incompetent adenovirus polyepitope vaccine is an excellent tool for the induction of a protective CTL response directed toward multiple LMP CTL epitopes restricted through common HLA class I alleles prevalent in different ethnic groups where EBV-associated malignancies are endemic.

Tyrosine 394 is phosphorylated in Alzheimer's paired helical filament tau and in fetal tau with c-Abl as the candidate tyrosine kinase

Tau is a major microtubule-associated protein of axons and is also the principal component of the paired helical filaments (PHFs) that comprise the neurofibrillary tangles found in Alzheimer's disease and other tauopathies. Besides phosphorylation of tau on serine and threonine residues in both normal tau and tau from neurofibrillary tangles, Tyr-18 was reported to be a site of phosphorylation by the Src-family kinase Fyn. We examined whether tyrosine residues other than Tyr-18 are phosphorylated in tau and whether other tyrosine kinases might phosphorylate tau. Using mass spectrometry, we positively identified phosphorylated Tyr-394 in PHF-tau from an Alzheimer brain and in human fetal brain tau. When wild-type human tau was transfected into fibroblasts or neuroblastoma cells, treatment with pervanadate caused tau to become phosphorylated on tyrosine by endogenous kinases. By replacing each of the five tyrosines in tau with phenylalanine, we identified Tyr-394 as the major site of tyrosine phosphorylation in tau. Tyrosine phosphorylation of tau was inhibited by PP2 (4-amino-5-(4-chlorophenyl-7-(t-butyl) pyrazolo[3,4-d] pyrimidine), which is known to inhibit Src-family kinases and c-Abl. Cotransfection of tau and kinases showed that Tyr-18 was the major site for Fyn phosphorylation, but Tyr-394 was the main residue for Abl. In vitro, Abl phosphorylated tau directly. Abl could be coprecipitated with tau and was present in pretangle neurons in brain sections from Alzheimer cases. These results show that phosphorylation of tau on Tyr-394 is a physiological event that is potentially part of a signal relay and suggest that Abl could have a pathogenic role in Alzheimer's disease.

Silencing of integrated human papillomavirus type 18 oncogene transcription in cells expressing SerpinB2

The serine protease inhibitor SerpinB2 (PAI-2), a major product of differentiating squamous epithelial cells, has recently been shown to bind and protect the retinoblastoma protein (Rb) from degradation. In human papillomavirus type 18 (HPV-18) -transformed epithelial cells the expression of the E6 and E7 oncoproteins is controlled by the HPV-18 upstream regulatory region (URR). Here we illustrate that PAI-2 expression in the HPV-18-transformed cervical carcinoma line HeLa resulted in the restoration of Rb expression, which led to the functional silencing of transcription from the HPV-18 URR. This caused loss of E7 protein expression and restoration of multiple E6- and E7-targeted host proteins, including p53, c-Myc, and c-Jun. Rb expression emerged as sufficient for the transcriptional repression of the URR, with repression mediated via the C/EB beta-YY1 binding site (URR 7709 to 7719). In contrast to HeLa cells, where the C/EBP beta-YY1 dimer binds this site, in PAI-2- and/or Rb-expressing cells the site was occupied by the dominant-negative C/EBP beta isoform liver-enriched transcriptional inhibitory protein (LIP). PAI-2 expression thus has a potent suppressive effect on HPV-18 oncogene transcription mediated by Rb and LIP, a finding with potential implications for prognosis and treatment of HPV-transformed lesions.

The Role of Ras Oncogene in Differentiation of 3T3-L1 Preadipocytes

Ras is a proto-oncogene that codes for a small GTPase and is responsible for linking several extracellular signals to intracellular mechanisms that involve cell growth, differentiation and cell-programmed death in normal and diseased cells. In all these processes, Ras has been extensively investigated. However, the role of Ras GTPases is still poorly understood during the differentiation of 3T3-L1 preadipocytes. In this study I investigated the role of the H-Ras defective mutant, Ras:G12V on the differentiation of 3T3-L1 preadipocytes. Preadipocytes were differentiated in vitro to adipocytes (fat cells) by adding an induction medium containing several factors including glucose and insulin. The formation of fat cells evidenced by the visualization of lipid drops as well as by quantifying the accumulation of Oil red O into lipid drops. To examine the role of Ras:G12V mutant, several selective mutations were introduced in order to determine the signaling transduction pathways (i.e., PI3(K)kinase and MAP(K)Kinase) responsible for the Ras-dependent adipogenesis. Cells expressing Ras:G12V mutant stimulated 3T3-L1 preadipocyte differentiation without he need for induction media, suggesting that Ras activation is an essential factor required for 3T3-L1 preadipocyte differentiation. Introduction of a second mutation on Ras:G12V (i.e., Ras:G12V;E37G), which blocks the activation of the MAPKinase pathway, strongly inhibited the 3T3-L1 preadipocyte differentiation. It is also important to note Ras:G12V:E37G double mutant does not inhibit the activation of the PI3kinase pathway. Other Ras double mutants (Ras:G12V;S35T, and V12G;C40Y) showed a modest inhibition of the 3T3-L1 preadipocyte differentiation. Taken together, these observations indicate that Ras plays a selective role in 3T3-L1 preadipocyte differentiation. Thus, understanding which specific pathway Ras employs during preadipocyte differentiation could clarify some of the uncertainties surrounding fat production.

Role of ABL Family Kinases in Breast Cancer

The ABL family of non-receptor tyrosine kinases, ABL1 (also known as c-ABL) and ABL2 (also known as Arg), links diverse extracellular stimuli to signaling pathways that control cell growth, survival, adhesion, migration and invasion. ABL tyrosine kinases play an oncogenic role in human leukemias. However, the role of ABL kinases in solid tumors including breast cancer progression and metastasis is just emerging.

To evaluate whether ABL family kinases are involved in breast cancer development and metastasis, we first analyzed genomic data from large-scale screen of breast cancer patients. We found that ABL kinases are up-regulated in invasive breast cancer patients and high expression of ABL kinases correlates with poor prognosis and early metastasis. Using xenograft mouse models combined with genetic and pharmacological approaches, we demonstrated that ABL kinases are required for regulating breast cancer progression and metastasis to the bone. Using next generation sequencing and bioinformatics analysis, we uncovered a critical role for ABL kinases in promoting multiple oncogenic pathways including TAZ and STAT5 signaling networks and the epithelial to mesenchymal transition (EMT). These findings revealed a role for ABL kinases in regulating breast cancer tumorigenesis and bone metastasis and provide a rationale for targeting breast tumors with ABL-specific inhibitors.

Investigation of the role of long non-coding RNAs in oncogene induced cellular senescence

Cellular senescence is a stable arrest of cell proliferation induced by several factors such as activated oncogenes, oxidative stress and shortening of telomeres. Senescence acts as a tumour suppression mechanism to halt the progression of cancer. However, senescence may also impact negatively upon tissue regeneration, thus contributing to the effects of ageing. The eukaryotic genome is controlled by various modes of transcriptional and translational regulation. Focus has therefore centred on the role of long non- coding RNAs (lncRNAs) in regulating the genome. Accordingly, understanding how lncRNAs function to regulate the senescent genome is integral to improving our knowledge and understanding of tumour suppression and ageing. Within this study, I set out to investigate the expression of lncRNAs’ expression within models of senescence. Through a custom expression array, I have shown that expression of multiple different lncRNAs is up-regulated and down regulated in IMR90 replicative senescent fibroblasts and oncogene-induced senescent melanocytes. LncRNA expression was determined to be specific to stable senescence-associated cell arrest and predominantly within the nucleus of senescent cells. In order to examine the function of lncRNA expression in senescence, I selected lncRNA transcript ENST0000430998 (lncRNA_98) to focus my investigations upon. LncRNA_98 was robustly upregulated within multiple models of senescence and efficiently depleted using anti-sense oligonucleotide technology. Characterisation and unbiased RNA-sequencing of lncRNA_98 deficient senescent cells highlighted a list of genes that are regulated by lncRNA_98 expression in senescent cells and may regulate aspects of the senescence program. Specifically, the formation of SAHF was impeded upon depletion of lncRNA_98 expression and levels of total pRB protein expression severely decreased. Validation and recapitulation of consequences of pRB depletion was confirmed through lncRNA_98 knock-out cells generated using CRISPR technology. Surprisingly, inhibition of ATM kinase functions permitted the restoration of pRB protein levels within lncRNA_98 deficient cells. I propose that lncRNA_98 antagonizes the ability of ATM kinase to downregulate pRB expression at a post-transcriptional level, thereby potentiating senescence. Furthermore, lncRNA expression was detected within fibroblasts of old individuals and visualised within senescent melanocytes in human benign nevi, a barrier to melanoma progression. Conversely, mining of 337 TCGA primary melanoma data sets highlighted that the lncRNA_98 gene and its expression was lost from a significant proportion of melanoma samples, consistent with lncRNA_98 having a tumour suppressor functions. The data presented in this study illustrates that lncRNA_98 expression has a regulatory role over pRB expression in senescence and may regulate aspects of tumourigenesis and ageing.

Clinical and molecular characterization of feline mammary carcinomas overexpressing HER2 proto-oncogene (FMC-HER2+) : new strategies for effective diagnostic and cancer therapy

Tese de Doutoramento em Ciências Veterinárias na Especialidade de Ciências Biológicas e Biomédicas

Prognostic Value Of Dna And Mrna E6/e7 Of Human Papillomavirus In The Evolution Of Cervical Intraepithelial Neoplasia Grade 2.

This study aimed at evaluating whether human papillomavirus (HPV) groups and E6/E7 mRNA of HPV 16, 18, 31, 33, and 45 are prognostic of cervical intraepithelial neoplasia (CIN) 2 outcome in women with a cervical smear showing a low-grade squamous intraepithelial lesion (LSIL). This cohort study included women with biopsy-confirmed CIN 2 who were followed up for 12 months, with cervical smear and colposcopy performed every three months. Women with a negative or low-risk HPV status showed 100% CIN 2 regression. The CIN 2 regression rates at the 12-month follow-up were 69.4% for women with alpha-9 HPV versus 91.7% for other HPV species or HPV-negative status (P < 0.05). For women with HPV 16, the CIN 2 regression rate at the 12-month follow-up was 61.4% versus 89.5% for other HPV types or HPV-negative status (P < 0.05). The CIN 2 regression rate was 68.3% for women who tested positive for HPV E6/E7 mRNA versus 82.0% for the negative results, but this difference was not statistically significant. The expectant management for women with biopsy-confirmed CIN 2 and previous cytological tests showing LSIL exhibited a very high rate of spontaneous regression. HPV 16 is associated with a higher CIN 2 progression rate than other HPV infections. HPV E6/E7 mRNA is not a prognostic marker of the CIN 2 clinical outcome, although this analysis cannot be considered conclusive. Given the small sample size, this study could be considered a pilot for future larger studies on the role of predictive markers of CIN 2 evolution.