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1.
Bioinformatics ; 36(1): 205-211, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31243428

RESUMO

MOTIVATION: The principal lines of research in MS/MS based Proteomics have been directed toward the molecular characterization of the proteins including their biological functions and their implications in human diseases. Recent advances in this field have also allowed the first attempts to apply these techniques to the clinical practice. Nowadays, the main progress in Computational Proteomics is based on the integration of genomic, transcriptomic and proteomic experimental data, what is known as Proteogenomics. This methodology is being especially useful for the discovery of new clinical biomarkers, small open reading frames and microproteins, although their validation is still challenging. RESULTS: We detected novel peptides following a proteogenomic workflow based on the MiTranscriptome human assembly and shotgun experiments. The annotation approach generated three custom databases with the corresponding peptides of known and novel transcripts of both protein coding genes and non-coding genes. In addition, we used a peptide detectability filter to improve the computational performance of the proteomic searches, the statistical analysis and the robustness of the results. These innovative additional filters are specially relevant when noisy next generation sequencing experiments are used to generate the databases. This resource, MiTPeptideDB, was validated using 43 cell lines for which RNA-Seq experiments and shotgun experiments were available. AVAILABILITY AND IMPLEMENTATION: MiTPeptideDB is available at http://bit.ly/MiTPeptideDB. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

2.
Bioinformatics ; 2019 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-31529040

RESUMO

SUMMARY: The protein detection and quantification using high-throughput proteomic technologies is still challenging due to the stochastic nature of the peptide selection in the mass spectrometer, the difficulties in the statistical analysis of the results and the presence of degenerated peptides. However, considering in the analysis only those peptides that could be detected by mass spectrometry (MS), also called proteotypic peptides, increases the accuracy of the results. Several approaches have been applied to predict peptide detectability based on the physicochemical properties of the peptides. In this manuscript we present DeepMSPeptide, a bioinformatic tool that uses a deep learning method to predict proteotypic peptides exclusively based on the peptide amino acid sequences. AVAILABILITY AND IMPLEMENTATION: DeepMSPeptide is available at https://github.com/vsegurar/DeepMSPeptide.

3.
Cancer Res ; 79(20): 5167-5180, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31387921

RESUMO

The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) datasets allow unprecedented gene expression analyses. Here, using these datasets, we performed pan-cancer and pan-tissue identification of coding and long noncoding RNA (lncRNA) transcripts differentially expressed in tumors and preferentially expressed in healthy tissues and/or tumors. Pan-cancer comparison of mRNAs and lncRNAs showed that lncRNAs were deregulated in a more tumor-specific manner. Given that lncRNAs are more tissue-specific than mRNAs, we identified healthy tissues that preferentially express lncRNAs upregulated in tumors and found that testis, brain, the digestive tract, and blood/spleen were the most prevalent. In addition, specific tumors also upregulate lncRNAs preferentially expressed in other tissues, generating a unique signature for each tumor type. Most tumors studied downregulated lncRNAs preferentially expressed in their tissue of origin, probably as a result of dedifferentiation. However, the same lncRNAs could be upregulated in other tumors, resulting in "bimorphic" transcripts. In hepatocellular carcinoma (HCC), the upregulated genes identified were expressed at higher levels in patients with worse prognosis. Some lncRNAs upregulated in HCC and preferentially expressed in healthy testis or brain were predicted to function as oncogenes and were significantly associated with higher tumor burden, and poor prognosis, suggesting their relevance in hepatocarcinogenesis and/or tumor evolution. Taken together, therapies targeting oncogenic lncRNAs should take into consideration the healthy tissue, where the lncRNAs are preferentially expressed, to predict and decrease unwanted secondary effects and increase potency. SIGNIFICANCE: Comprehensive analysis of coding and noncoding genes expressed in different tumors and normal tissues, which should be taken into account to predict side effects from potential coding and noncoding gene-targeting therapies.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/20/5167/F1.large.jpg.

4.
Front Aging Neurosci ; 11: 149, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31281249

RESUMO

The discouraging results with therapies for Alzheimer's disease (AD) in clinical trials, highlights the urgent need to adopt new approaches. Like other complex diseases, it is becoming clear that AD therapies should focus on the simultaneous modulation of several targets implicated in the disease. Recently, using reference compounds and the first-in class CM-414, we demonstrated that the simultaneous inhibition of histone deacetylases [class I histone deacetylases (HDACs) and HDAC6] and phosphodiesterase 5 (PDE5) has a synergistic therapeutic effect in AD models. To identify the best inhibitory balance of HDAC isoforms and PDEs that provides a safe and efficient therapy to combat AD, we tested the compound CM-695 in the Tg2576 mouse model of this disease. CM-695 selectively inhibits HDAC6 over class I HDAC isoforms, which largely overcomes the toxicity associated with HDAC class 1 inhibition. Furthermore, CM-695 inhibits PDE9, which is expressed strongly in the brain and has been proposed as a therapeutic target for AD. Chronic treatment of aged Tg2576 mice with CM-695 ameliorates memory impairment and diminishes brain Aß, although its therapeutic effect was no longer apparent 4 weeks after the treatment was interrupted. An increase in the presence of 78-KDa glucose regulated protein (GRP78) and heat shock protein 70 (Hsp70) chaperones may underlie the therapeutic effect of CM-695. In summary, chronic treatment with CM-695 appears to reverse the AD phenotype in a safe and effective manner. Taking into account that AD is a multifactorial disorder, the multimodal action of these compounds and the different events they affect may open new avenues to combat AD.

5.
Int J Cancer ; 145(7): 1991-2001, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-30848481

RESUMO

Sunitinib is one of the most widely used targeted therapeutics for renal cell carcinoma (RCC), but acquired resistance against targeted therapies remains a major clinical challenge. To dissect mechanisms of acquired resistance and unravel reliable predictive biomarkers for sunitinib in RCC, we sequenced the exons of 409 tumor-suppressor genes and oncogenes in paired tumor samples from an RCC patient, obtained at baseline and after development of acquired resistance to sunitinib. From newly arising mutations, we selected, using in silico prediction models, six predicted to be deleterious, located in G6PD, LRP1B, SETD2, TET2, SYNE1, and DCC. Consistently, immunoblotting analysis of lysates derived from sunitinib-desensitized RCC cells and their parental counterparts showed marked differences in the levels and expression pattern of the proteins encoded by these genes. Our further analysis demonstrates essential roles for these proteins in mediating sunitinib cytotoxicity and shows that their loss of function renders tumor cells resistant to sunitinib in vitro and in vivo. Finally, sunitinib resistance induced by continuous exposure or by inhibition of the six proteins was overcome by treatment with cabozantinib or a low-dose combination of lenvatinib and everolimus. Collectively, our results unravel novel markers of acquired resistance to sunitinib and clinically relevant approaches for overcoming this resistance in RCC.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Renais/genética , Resistencia a Medicamentos Antineoplásicos , Neoplasias Renais/genética , Mutação , Animais , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Éxons , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Renais/metabolismo , Camundongos , Transplante de Neoplasias , Análise de Sequência de DNA , Sunitinibe
6.
Expert Rev Proteomics ; 16(3): 267-275, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30654666

RESUMO

INTRODUCTION: The technological and scientific progress performed in the Human Proteome Project (HPP) has provided to the scientific community a new set of experimental and bioinformatic methods in the challenging field of shotgun and SRM/MRM-based Proteomics. The requirements for a protein to be considered experimentally validated are now well-established, and the information about the human proteome is available in the neXtProt database, while targeted proteomic assays are stored in SRMAtlas. However, the study of the missing proteins continues being an outstanding issue. Areas covered: This review is focused on the implementation of proteogenomic methods designed to improve the detection and validation of the missing proteins. The evolution of the methodological strategies based on the combination of different omic technologies and the use of huge publicly available datasets is shown taking the Chromosome 16 Consortium as reference. Expert commentary: Proteogenomics and other strategies of data analysis implemented within the C-HPP initiative could be used as guidance to complete in a near future the catalog of the human proteins. Besides, in the next years, we will probably witness their use in the B/D-HPP initiative to go a step forward on the implications of the proteins in the human biology and disease.

7.
Clin Cancer Res ; 25(10): 3176-3187, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30692097

RESUMO

PURPOSE: Knowledge about the mechanism of action (MoA) of monoclonal antibodies (mAb) is required to understand which patients with multiple myeloma (MM) benefit the most from a given mAb, alone or in combination therapy. Although there is considerable research about daratumumab, knowledge about other anti-CD38 mAbs remains scarce. EXPERIMENTAL DESIGN: We performed a comprehensive analysis of the MoA of isatuximab. RESULTS: Isatuximab induces internalization of CD38 but not its significant release from MM cell surface. In addition, we uncovered an association between levels of CD38 expression and different MoA: (i) Isatuximab was unable to induce direct apoptosis on MM cells with CD38 levels closer to those in patients with MM, (ii) isatuximab sensitized CD38hi MM cells to bortezomib plus dexamethasone in the presence of stroma, (iii) antibody-dependent cellular cytotoxicity (ADCC) was triggered by CD38lo and CD38hi tumor plasma cells (PC), (iv) antibody-dependent cellular phagocytosis (ADCP) was triggered only by CD38hi MM cells, whereas (v) complement-dependent cytotoxicity could be triggered in less than half of the patient samples (those with elevated levels of CD38). Furthermore, we showed that isatuximab depletes CD38hi B-lymphocyte precursors and natural killer (NK) lymphocytes ex vivo-the latter through activation followed by exhaustion and eventually phagocytosis. CONCLUSIONS: This study provides a framework to understand response determinants in patients treated with isatuximab based on the number of MoA triggered by CD38 levels of expression, and for the design of effective combinations aimed at capitalizing disrupted tumor-stroma cell protection, augmenting NK lymphocyte-mediated ADCC, or facilitating ADCP in CD38lo MM patients.See related commentary by Malavasi and Faini, p. 2946.


Assuntos
Mieloma Múltiplo/imunologia , ADP-Ribosil Ciclase 1/imunologia , Anticorpos Monoclonais , Humanos
8.
Haematologica ; 104(8): 1572-1579, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30655376

RESUMO

In this study we interrogated the DNA methylome of myelofibrosis patients using high-density DNA methylation arrays. We detected 35,215 differentially methylated CpG, corresponding to 10,253 genes, between myelofibrosis patients and healthy controls. These changes were present both in primary and secondary myelofibrosis, which showed no differences between them. Remarkably, most differentially methylated CpG were located outside gene promoter regions and showed significant association with enhancer regions. This aberrant enhancer hypermethylation was negatively correlated with the expression of 27 genes in the myelofibrosis cohort. Of these, we focused on the ZFP36L1 gene and validated its decreased expression and enhancer DNA hypermethylation in an independent cohort of patients and myeloid cell-lines. In vitro reporter assay and 5'-azacitidine treatment confirmed the functional relevance of hyper-methylation of ZFP36L1 enhancer. Furthermore, in vitro rescue of ZFP36L1 expression had an impact on cell proliferation and induced apoptosis in SET-2 cell line indicating a possible role of ZFP36L1 as a tumor suppressor gene in myelofibrosis. Collectively, we describe the DNA methylation profile of myelofibrosis, identifying extensive changes in enhancer elements and revealing ZFP36L1 as a novel candidate tumor suppressor gene.

10.
Oncotarget ; 9(16): 12842-12852, 2018 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-29560114

RESUMO

Long Non-Coding RNAs (lncRNAs) are functional RNAs longer than 200 nucleotides in length. Several lncRNAs are involved in cell proliferation and are deregulated in several human tumors. Few lncRNAs have been described to play a role in Acute Lymphoblastic Leukemia (ALL). In this study, we carried out a genome wide lncRNA expression profiling in ALL samples and peripheral blood samples obtained from healthy donors. We detected 43 lncRNAs that were aberrantly expressed in ALL. Interestingly, among them, linc-PINT showed a significant downregulation in T and B-ALL. Re-expression of linc-PINT in ALL cells induced inhibition of leukemic cell growth that was associated with apoptosis induction and cell cycle arrest in G2/M phase. linc-PINT induced the transcription of HMOX1 which reduced the viability of ALL cells. Intriguingly, we observed that treatment with anti-tumoral epigenetic drugs like LBH-589 (Panobinostat) and Curcumin induced the expression of linc-PINT and HMOX1 in ALL. These results indicate that the downregulation of linc-PINT plays a relevant role in the pathogenesis of ALL, and linc-PINT re-expression may be one of the mechanisms exerted by epigenetic drugs to reduce cell proliferation in ALL.

11.
J Pathol ; 245(1): 61-73, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29464716

RESUMO

The increased risk of Richter transformation (RT) in patients with chronic lymphocytic leukaemia (CLL) due to Epstein-Barr virus (EBV) reactivation during immunosuppressive therapy with fludarabine other targeted agents remains controversial. Among 31 RT cases classified as diffuse large B-cell lymphoma (DLBCL), seven (23%) showed EBV expression. In contrast to EBV- tumours, EBV+ DLBCLs derived predominantly from IGVH-hypermutated CLL, and they also showed CLL-unrelated IGVH sequences more frequently. Intriguingly, despite having different cellular origins, clonally related and unrelated EBV+ DLBCLs shared a previous history of immunosuppressive chemo-immunotherapy, a non-germinal centre DLBCL phenotype, EBV latency programme type II or III, and very short survival. These data suggested that EBV reactivation during therapy-related immunosuppression can transform either CLL cells or non-tumoural B lymphocytes into EBV+ DLBCL. To investigate this hypothesis, xenogeneic transplantation of blood cells from 31 patients with CLL and monoclonal B-cell lymphocytosis (MBL) was performed in Rag2-/- IL2γc-/- mice. Remarkably, the recipients' impaired immunosurveillance favoured the spontaneous outgrowth of EBV+ B-cell clones from 95% of CLL and 64% of MBL patients samples, but not from healthy donors. Eventually, these cells generated monoclonal tumours (mostly CLL-unrelated but also CLL-related), recapitulating the principal features of EBV+ DLBCL in patients. Accordingly, clonally related and unrelated EBV+ DLBCL xenografts showed indistinguishable cellular, virological and molecular features, and synergistically responded to combined inhibition of EBV replication with ganciclovir and B-cell receptor signalling with ibrutinib in vivo. Our study underscores the risk of RT driven by EBV in CLL patients receiving immunosuppressive therapies, and provides the scientific rationale for testing ganciclovir and ibrutinib in EBV+ DLBCL. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Assuntos
Transformação Celular Neoplásica/efeitos dos fármacos , Herpesvirus Humano 4/efeitos dos fármacos , Imunossupressores/farmacologia , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Adulto , Idoso , Linfócitos B/efeitos dos fármacos , Linfócitos B/patologia , Transformação Celular Neoplásica/patologia , Infecções por Vírus Epstein-Barr/tratamento farmacológico , Infecções por Vírus Epstein-Barr/patologia , Feminino , Herpesvirus Humano 4/genética , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/genética , Masculino , Pessoa de Meia-Idade
12.
Proteomes ; 6(1)2018 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-29401756

RESUMO

Monocytes are bone marrow-derived leukocytes that are part of the innate immune system. Monocytes are divided into three subsets: classical, intermediate and non-classical, which can be differentiated by their expression of some surface antigens, mainly CD14 and CD16. These cells are key players in the inflammation process underlying the mechanism of many diseases. Thus, the molecular characterization of these cells may provide very useful information for understanding their biology in health and disease. We performed a multicentric proteomic study with pure classical and non-classical populations derived from 12 healthy donors. The robust workflow used provided reproducible results among the five participating laboratories. Over 5000 proteins were identified, and about half of them were quantified using a spectral counting approach. The results represent the protein abundance catalogue of pure classical and enriched non-classical blood peripheral monocytes, and could serve as a reference dataset of the healthy population. The functional analysis of the differences between cell subsets supports the consensus roles assigned to human monocytes.

13.
Nucleic Acids Res ; 46(3): 1345-1361, 2018 02 16.
Artigo em Inglês | MEDLINE | ID: mdl-29309682

RESUMO

RNA-binding proteins (RBPs) are essential to fine-tune gene expression. RBPs containing the cold-shock domain are RNA chaperones that have been extensively studied. However, the RNA targets and specific functions for many of them remain elusive. Here, combining comparative proteomics and RBP-immunoprecipitation-microarray profiling, we have determined the regulon of the RNA chaperone CspA of Staphylococcus aureus. Functional analysis revealed that proteins involved in carbohydrate and ribonucleotide metabolism, stress response and virulence gene expression were affected by cspA deletion. Stress-associated phenotypes such as increased bacterial aggregation and diminished resistance to oxidative-stress stood out. Integration of the proteome and targetome showed that CspA post-transcriptionally modulates both positively and negatively the expression of its targets, denoting additional functions to the previously proposed translation enhancement. One of these repressed targets was its own mRNA, indicating the presence of a negative post-transcriptional feedback loop. CspA bound the 5'UTR of its own mRNA disrupting a hairpin, which was previously described as an RNase III target. Thus, deletion of the cspA 5'UTR abrogated mRNA processing and auto-regulation. We propose that CspA interacts through a U-rich motif, which is located at the RNase III cleavage site, portraying CspA as a putative RNase III-antagonist.


Assuntos
Proteínas de Bactérias/genética , Retroalimentação Fisiológica , Regulação Bacteriana da Expressão Gênica , Proteoma/genética , Regulon , Ribonuclease III/genética , Staphylococcus aureus/genética , Regiões 5' não Traduzidas , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Pareamento de Bases , Sítios de Ligação , Metabolismo dos Carboidratos/genética , Deleção de Genes , Modelos Moleculares , Conformação de Ácido Nucleico , Ligação Proteica , Estrutura Secundária de Proteína , Proteoma/metabolismo , RNA Bacteriano , Ribonuclease III/química , Ribonuclease III/metabolismo , Staphylococcus aureus/metabolismo , Staphylococcus aureus/patogenicidade , Estresse Fisiológico/genética , Virulência
14.
Epigenomics ; 10(1): 91-103, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29172706

RESUMO

AIM: To analyze whether preterm newborns show differences in methylation patterns in comparison to full-term newborns in white blood cells. PATIENTS & METHODS: Anthropometrical, biochemical features and methylation levels of preterm newborns (n = 24) and full-term newborns (n = 22) recruited in La Paz University Hospital (Spain) were assessed at 12 months of gestational age, whereas Bayley Scale of Infant Development was evaluated at 24/36 months. RESULTS: From all the statistically significant CpGs, methylation levels of cg00997378 (SLC6A3 gene) showed the highest differences (p < 0.0001), being associated with prematurity risk factors. CONCLUSION: SLC6A3 methylation, previously related to attention-deficit/hyperactivity disorder, neuronal function and behavior, might be a potential epigenetic biomarker with value in the early diagnosis and management of neurodevelopmental diseases in newborns.


Assuntos
Metilação de DNA , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Sistema Nervoso/crescimento & desenvolvimento , Ilhas de CpG , Feminino , Humanos , Recém-Nascido Prematuro , Leucócitos/metabolismo , Masculino
15.
PLoS One ; 12(12): e0190275, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29281720

RESUMO

The combination of defined factors with small molecules targeting epigenetic factors is a strategy that has been shown to enhance optimal derivation of iPSCs and could be used for disease modelling, high throughput screenings and/or regenerative medicine applications. In this study, we showed that a new first-in-class reversible dual G9a/DNMT1 inhibitor compound (CM272) improves the efficiency of human cell reprogramming and iPSC generation from primary cells of healthy donors and patient samples, using both integrative and non-integrative methods. Moreover, CM272 facilitates the generation of human iPSC with only two factors allowing the removal of the most potent oncogenic factor cMYC. Furthermore, we demonstrated that mechanistically, treatment with CM272 induces heterochromatin relaxation, facilitates the engagement of OCT4 and SOX2 transcription factors to OSKM refractory binding regions that are required for iPSC establishment, and enhances mesenchymal to epithelial transition during the early phase of cell reprogramming. Thus, the use of this new G9a/DNMT reversible dual inhibitor compound may represent an interesting alternative for improving cell reprogramming and human iPSC derivation for many different applications while providing interesting insights into reprogramming mechanisms.


Assuntos
Reprogramação Celular , Genoma Humano , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Células-Tronco Pluripotentes Induzidas/citologia , Proteínas Repressoras/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Células Cultivadas , Antígenos de Histocompatibilidade , Humanos , Reação em Cadeia da Polimerase em Tempo Real
16.
Oncotarget ; 8(41): 69663-69679, 2017 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-29050232

RESUMO

Alzheimer´s disease (AD) is characterized by progressive dementia, initially presenting olfactory dysfunction. Despite the olfactory bulb (OB) is the first central structure of the olfactory pathway, we lack a complete molecular characterization of the transcriptional events that occurs in this olfactory area during AD progression. To address this gap in knowledge, we have assessed the genome-wide expression in postmortem OBs from subjects with varying degree of AD pathology. A stage-dependent deregulation of specific pathways was observed, revealing transmembrane transport, and neuroinflammation as part of the functional modules that are disrupted across AD grading. Potential drivers of neurodegeneration predicted by network-driven transcriptomics were monitored across different types of dementia, including progressive supranuclear palsy (PSP), mixed dementia, and frontotemporal lobar degeneration (FTLD). Epidermal growth factor receptor (EGFR) expression was significantly increased in the OB of AD and mixed dementia subjects. Moreover, a significant increment in the activation of signal transducer and activator of transcription 3 (STAT3) was exclusively detected in advanced AD stages, whereas total STAT3 levels were specifically overexpressed in mixed dementia. Furthermore, transcription factors deregulated in the OB of mixed dementia subjects such as cAMP Responsive Element Binding Protein 1 (CREB1) and AP-1 Transcription Factor Subunit (c-Jun) were not differentially modulated at olfactory level across AD grading. On the other hand, olfactory expression of this signal transducer panel was unchanged in PSP and FTLD subjects. Taken together, this study unveils cross-disease similarities and differences for specific signal transducers, providing mechanistic clues to the intriguing divergence of AD pathology across proteinopathies.

17.
J Proteome Res ; 16(12): 4374-4390, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-28960077

RESUMO

The Human Proteome Project (HPP) aims deciphering the complete map of the human proteome. In the past few years, significant efforts of the HPP teams have been dedicated to the experimental detection of the missing proteins, which lack reliable mass spectrometry evidence of their existence. In this endeavor, an in depth analysis of shotgun experiments might represent a valuable resource to select a biological matrix in design validation experiments. In this work, we used all the proteomic experiments from the NCI60 cell lines and applied an integrative approach based on the results obtained from Comet, Mascot, OMSSA, and X!Tandem. This workflow benefits from the complementarity of these search engines to increase the proteome coverage. Five missing proteins C-HPP guidelines compliant were identified, although further validation is needed. Moreover, 165 missing proteins were detected with only one unique peptide, and their functional analysis supported their participation in cellular pathways as was also proposed in other studies. Finally, we performed a combined analysis of the gene expression levels and the proteomic identifications from the common cell lines between the NCI60 and the CCLE project to suggest alternatives for further validation of missing protein observations.


Assuntos
Proteoma/análise , Proteômica/métodos , Ferramenta de Busca , Linhagem Celular Tumoral , Humanos , Bases de Conhecimento , Proteínas/análise , Software
18.
Nat Commun ; 8: 15424, 2017 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-28548080

RESUMO

The indisputable role of epigenetics in cancer and the fact that epigenetic alterations can be reversed have favoured development of epigenetic drugs. In this study, we design and synthesize potent novel, selective and reversible chemical probes that simultaneously inhibit the G9a and DNMTs methyltransferase activity. In vitro treatment of haematological neoplasia (acute myeloid leukaemia-AML, acute lymphoblastic leukaemia-ALL and diffuse large B-cell lymphoma-DLBCL) with the lead compound CM-272, inhibits cell proliferation and promotes apoptosis, inducing interferon-stimulated genes and immunogenic cell death. CM-272 significantly prolongs survival of AML, ALL and DLBCL xenogeneic models. Our results represent the discovery of first-in-class dual inhibitors of G9a/DNMTs and establish this chemical series as a promising therapeutic tool for unmet needs in haematological tumours.


Assuntos
Antineoplásicos/farmacologia , Metilases de Modificação do DNA/antagonistas & inibidores , Desenho de Drogas , Inibidores Enzimáticos/farmacologia , Neoplasias Hematológicas/tratamento farmacológico , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Animais , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/genética , Apoptose/imunologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Cristalografia por Raios X , Metilases de Modificação do DNA/química , Metilases de Modificação do DNA/genética , Metilases de Modificação do DNA/metabolismo , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/uso terapêutico , Epigênese Genética/efeitos dos fármacos , Feminino , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/imunologia , Neoplasias Hematológicas/mortalidade , Antígenos de Histocompatibilidade/química , Antígenos de Histocompatibilidade/genética , Antígenos de Histocompatibilidade/metabolismo , Histona-Lisina N-Metiltransferase/química , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Interferons/imunologia , Interferons/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Microssomos Hepáticos , Simulação de Acoplamento Molecular , Análise de Sobrevida , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
19.
J Hepatol ; 67(4): 669-679, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28527664

RESUMO

BACKGROUND & AIMS: Studying hepatitis delta virus (HDV) and developing new treatments is hampered by the limited availability of small animal models. Herein, a description of a robust mouse model of HDV infection that mimics several important characteristics of the human disease is presented. METHODS: HDV and hepatitis B virus (HBV) replication competent genomes were delivered to the mouse liver using adeno-associated viruses (AAV; AAV-HDV and AAV-HBV). Viral load, antigen expression and genomes were quantified at different time points after AAV injection. Furthermore, liver pathology, genome editing, and the activation of the innate immune response were evaluated. RESULTS: AAV-HDV infection initiated HDV replication in mouse hepatocytes. Genome editing was confirmed by the presence of small and large HDV antigens and sequencing. Viral replication was detected for 45days, even after the AAV-HDV vector had almost disappeared. In the presence of HBV, HDV infectious particles were detected in serum. Furthermore, as observed in patients, co-infection was associated with the reduction of HBV antigen expression and the onset of liver damage that included the alteration of genes involved in the development of liver pathologies. HDV replication induced a sustained type I interferon response, which was significantly reduced in immunodeficient mice and almost absent in mitochondrial antiviral signaling protein (MAVS)-deficient mice. CONCLUSION: The animal model described here reproduces important characteristics of human HDV infection and provides a valuable tool for characterizing the viral infection and for developing new treatments. Furthermore, MAVS was identified as a main player in HDV detection and adaptive immunity was found to be involved in the amplification of the innate immune response. Lay summary: Co-infection with hepatitis B and D virus (HBV and HDV, respectively) often causes a more severe disease condition than HBV alone. Gaining more insight into HDV and developing new treatments is hampered by limited availability of adequate immune competent small animal models and new ones are needed. Here, a mouse model of HDV infection is described, which mimics several important characteristics of the human disease, such as the initiation and maintenance of replication in murine hepatocytes, genome editing and, in the presence of HBV, generation of infectious particles. Lastly, the involvement of an adaptive immunity and the intracellular signaling molecule MAVS in mounting a strong and lasting innate response was shown. Thus, our model serves as a useful tool for the investigation of HDV biology and new treatments.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/imunologia , Hepatite D/imunologia , Interferon beta/biossíntese , Imunidade Adaptativa , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Linhagem Celular , Coinfecção/imunologia , Coinfecção/patologia , Coinfecção/virologia , Dependovirus/genética , Modelos Animais de Doenças , Genoma Viral , Hepatite B/complicações , Hepatite B/imunologia , Hepatite B/virologia , Antígenos da Hepatite B/metabolismo , Vírus da Hepatite B/genética , Vírus da Hepatite B/imunologia , Hepatite D/complicações , Hepatite D/virologia , Vírus Delta da Hepatite/genética , Vírus Delta da Hepatite/imunologia , Vírus Delta da Hepatite/fisiologia , Antígenos da Hepatite delta/metabolismo , Humanos , Imunidade Inata , Fígado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Modelos Imunológicos , Transdução de Sinais/imunologia , Replicação Viral
20.
Cancer Treat Rev ; 53: 79-97, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28088073

RESUMO

The discovery of reliable biomarkers to predict efficacy and toxicity of anticancer drugs remains one of the key challenges in cancer research. Despite its relevance, no efficient study designs to identify promising candidate biomarkers have been established. This has led to the proliferation of a myriad of exploratory studies using dissimilar strategies, most of which fail to identify any promising targets and are seldom validated. The lack of a proper methodology also determines that many anti-cancer drugs are developed below their potential, due to failure to identify predictive biomarkers. While some drugs will be systematically administered to many patients who will not benefit from them, leading to unnecessary toxicities and costs, others will never reach registration due to our inability to identify the specific patient population in which they are active. Despite these drawbacks, a limited number of outstanding predictive biomarkers have been successfully identified and validated, and have changed the standard practice of oncology. In this manuscript, a multidisciplinary panel reviews how those key biomarkers were identified and, based on those experiences, proposes a methodological framework-the DESIGN guidelines-to standardize the clinical design of biomarker identification studies and to develop future research in this pivotal field.


Assuntos
Biomarcadores Tumorais/análise , Pesquisa Biomédica/métodos , Quinase do Linfoma Anaplásico , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Pesquisa Biomédica/normas , Estudos Clínicos como Assunto , Ensaios Clínicos como Assunto , Receptores ErbB/genética , Humanos , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-kit/genética , Receptores Proteína Tirosina Quinases/genética , Receptor ErbB-2/análise , Proteínas ras/genética
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