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1.
Cancer Cell ; 42(2): 225-237.e5, 2024 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-38278149

RESUMO

Small cell lung cancer (SCLC) is an aggressive malignancy composed of distinct transcriptional subtypes, but implementing subtyping in the clinic has remained challenging, particularly due to limited tissue availability. Given the known epigenetic regulation of critical SCLC transcriptional programs, we hypothesized that subtype-specific patterns of DNA methylation could be detected in tumor or blood from SCLC patients. Using genomic-wide reduced-representation bisulfite sequencing (RRBS) in two cohorts totaling 179 SCLC patients and using machine learning approaches, we report a highly accurate DNA methylation-based classifier (SCLC-DMC) that can distinguish SCLC subtypes. We further adjust the classifier for circulating-free DNA (cfDNA) to subtype SCLC from plasma. Using the cfDNA classifier (cfDMC), we demonstrate that SCLC phenotypes can evolve during disease progression, highlighting the need for longitudinal tracking of SCLC during clinical treatment. These data establish that tumor and cfDNA methylation can be used to identify SCLC subtypes and might guide precision SCLC therapy.


Assuntos
Ácidos Nucleicos Livres , Neoplasias Pulmonares , Carcinoma de Pequenas Células do Pulmão , Humanos , Carcinoma de Pequenas Células do Pulmão/genética , Carcinoma de Pequenas Células do Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Metilação de DNA , Ácidos Nucleicos Livres/genética , Epigênese Genética , Biomarcadores Tumorais/genética
3.
NAR Cancer ; 5(2): zcad022, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37206360

RESUMO

Maintenance of genomic methylation patterns at DNA replication forks by DNMT1 is the key to faithful mitotic inheritance. DNMT1 is often overexpressed in cancer cells and the DNA hypomethylating agents azacytidine and decitabine are currently used in the treatment of hematologic malignancies. However, the toxicity of these cytidine analogs and their ineffectiveness in treating solid tumors have limited wider clinical use. GSK-3484862 is a newly-developed, dicyanopyridine containing, non-nucleoside DNMT1-selective inhibitor with low cellular toxicity. Here, we show that GSK-3484862 targets DNMT1 for protein degradation in both cancer cell lines and murine embryonic stem cells (mESCs). DNMT1 depletion was rapid, taking effect within hours following GSK-3484862 treatment, leading to global hypomethylation. Inhibitor-induced DNMT1 degradation was proteasome-dependent, with no discernible loss of DNMT1 mRNA. In mESCs, GSK-3484862-induced Dnmt1 degradation requires the Dnmt1 accessory factor Uhrf1 and its E3 ubiquitin ligase activity. We also show that Dnmt1 depletion and DNA hypomethylation induced by the compound are reversible after its removal. Together, these results indicate that this DNMT1-selective degrader/inhibitor will be a valuable tool for dissecting coordinated events linking DNA methylation to gene expression and identifying downstream effectors that ultimately regulate cellular response to altered DNA methylation patterns in a tissue/cell-specific manner.

4.
Cancers (Basel) ; 14(13)2022 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-35805010

RESUMO

The aggressive variant prostate cancer molecular profile (AVPC-m), composed of combined defects in TP53, RB1 and PTEN, characterizes a subset of prostate cancers linked to androgen indifference and platinum sensitivity. To contribute to the optimization of the AVPC-m assessment for inclusion in prospective clinical trials, we investigated the status of the AVPC-m components in 28 patient tumor-derived xenografts (PDXs) developed at MDACC. We subjected single formalin-fixed, paraffin-embedded (FFPE) blocks from each PDX to immunohistochemistry (IHC), targeted next-generation genomic sequencing (NGS) and Clariom-S Affymetrix human microarray expression profiling. Standard validated IHC assays and a 10% labeling index cutoff resulted in high reproducibility across three separate laboratories and three independent readers for all tumor suppressors, as well as strong correlations with loss-of-function transcriptional scores (LOF-TS). Adding intensity assessment to labeling indices strengthened the association between IHC results and LOF-TS for TP53 and RB1, but not for PTEN. For TP53, genomic alterations determined by NGS had slightly higher agreement scores with LOF-TS than aberrant IHC, while for RB1 and PTEN, NGS and IHC determinations resulted in similar agreement scores with LOF-TS. Nonetheless, our results indicate that the AVPC-m components can be assessed reproducibly by IHC using various widely available standardized assays.

5.
Cancers (Basel) ; 14(3)2022 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-35158923

RESUMO

To be dormant or not depends on the origin and nature of both the cell and its niche. Similar to other cancer hallmarks, dormancy is ingrained with stemness, and stemness is embedded within dormancy. After all, cancer dormancy is dependent on multiple factors such as cell cycle arrest, metabolic inactivity, and the microenvironment. It is the net results and sum effects of a myriad of cellular interactions, interconnections, and interplays. When we unite all cancer networks and integrate all cancer hallmarks, we practice and preach a unified theory of cancer. From this perspective, we review cancer dormancy in the context of a stem cell theory of cancer. We revisit the seed and soil hypothesis of cancer. We reexamine its implications in both primary tumors and metastatic lesions. We reassess its roles in cell cycle arrest, metabolic inactivity, and stemness property. Cancer dormancy is particularly revealing when it informs us about the mysteries of late relapse, prolonged remission, and second malignancy. It is paradoxically rewarding when it delivers us the promises and power of cancer prevention and maintenance therapy in patient care.

6.
Nat Genet ; 53(10): 1456-1468, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34594038

RESUMO

Glioma intratumoral heterogeneity enables adaptation to challenging microenvironments and contributes to therapeutic resistance. We integrated 914 single-cell DNA methylomes, 55,284 single-cell transcriptomes and bulk multi-omic profiles across 11 adult IDH mutant or IDH wild-type gliomas to delineate sources of intratumoral heterogeneity. We showed that local DNA methylation disorder is associated with cell-cell DNA methylation differences, is elevated in more aggressive tumors, links with transcriptional disruption and is altered during the environmental stress response. Glioma cells under in vitro hypoxic and irradiation stress increased local DNA methylation disorder and shifted cell states. We identified a positive association between genetic and epigenetic instability that was supported in bulk longitudinally collected DNA methylation data. Increased DNA methylation disorder associated with accelerated disease progression and recurrently selected DNA methylation changes were enriched for environmental stress response pathways. Our work identified an epigenetically facilitated adaptive stress response process and highlights the importance of epigenetic heterogeneity in shaping therapeutic outcomes.


Assuntos
Neoplasias Encefálicas/genética , Plasticidade Celular/genética , Epigênese Genética , Glioma/genética , Análise de Célula Única , Estresse Fisiológico/genética , Evolução Clonal , Variações do Número de Cópias de DNA/genética , Metilação de DNA/genética , Regulação Neoplásica da Expressão Gênica , Heterogeneidade Genética , Genoma Humano , Humanos , Mutação/genética , Filogenia , Regiões Promotoras Genéticas/genética , Microambiente Tumoral/genética
7.
Hum Mol Genet ; 31(2): 262-274, 2021 12 27.
Artigo em Inglês | MEDLINE | ID: mdl-34432028

RESUMO

Myotonic dystrophy type 1 (DM1) is a complex disease with a wide spectrum of symptoms. The exact relationship between mutant CTG repeat expansion size and clinical outcome remains unclear. DM1 congenital patients (CDM) inherit the largest expanded alleles, which are associated with abnormal and increased DNA methylation flanking the CTG repeat. However, DNA methylation at the DMPK locus remains understudied. Its relationship to DM1 clinical subtypes, expansion size and age-at-onset is not yet completely understood. Using pyrosequencing-based methylation analysis on 225 blood DNA samples from Costa Rican DM1 patients, we determined that the size of the estimated progenitor allele length (ePAL) is not only a good discriminator between CDM and non-CDM cases (with an estimated threshold at 653 CTG repeats), but also for all DM1 clinical subtypes. Secondly, increased methylation at both CTCF sites upstream and downstream of the expansion was almost exclusively present in CDM cases. Thirdly, levels of abnormal methylation were associated with clinical subtype, age and ePAL, with strong correlations between these variables. Fourthly, both ePAL and the intergenerational expansion size were significantly associated with methylation status. Finally, methylation status was associated with ePAL and maternal inheritance, with almost exclusively maternal transmission of CDM. In conclusion, increased DNA methylation at the CTCF sites flanking the DM1 expansion could be linked to ePAL, and both increased methylation and the ePAL could be considered biomarkers for the CDM phenotype.


Assuntos
Distrofia Miotônica , Alelos , Fator de Ligação a CCCTC , Metilação de DNA/genética , Humanos , Distrofia Miotônica/genética , Miotonina Proteína Quinase/genética , Expansão das Repetições de Trinucleotídeos/genética
8.
Nat Commun ; 12(1): 687, 2021 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-33514726

RESUMO

The evolution of DNA methylome and methylation intra-tumor heterogeneity (ITH) during early carcinogenesis of lung adenocarcinoma has not been systematically studied. We perform reduced representation bisulfite sequencing of invasive lung adenocarcinoma and its precursors, atypical adenomatous hyperplasia, adenocarcinoma in situ and minimally invasive adenocarcinoma. We observe gradual increase of methylation aberrations and significantly higher level of methylation ITH in later-stage lesions. The phylogenetic patterns inferred from methylation aberrations resemble those based on somatic mutations suggesting parallel methylation and genetic evolution. De-convolution reveal higher ratio of T regulatory cells (Tregs) versus CD8 + T cells in later-stage diseases, implying progressive immunosuppression with neoplastic progression. Furthermore, increased global hypomethylation is associated with higher mutation burden, copy number variation burden and AI burden as well as higher Treg/CD8 ratio, highlighting the potential impact of methylation on chromosomal instability, mutagenesis and tumor immune microenvironment during early carcinogenesis of lung adenocarcinomas.


Assuntos
Adenocarcinoma de Pulmão/genética , Metilação de DNA/genética , Epigenoma/genética , Neoplasias Pulmonares/genética , Lesões Pré-Cancerosas/genética , Adenocarcinoma de Pulmão/mortalidade , Adenocarcinoma de Pulmão/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinogênese/genética , Instabilidade Cromossômica , Variações do Número de Cópias de DNA , Progressão da Doença , Feminino , Heterogeneidade Genética , Humanos , Hiperplasia/genética , Hiperplasia/patologia , Pulmão/patologia , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Mutagênese , Taxa de Mutação , Lesões Pré-Cancerosas/patologia , Análise de Sobrevida , Microambiente Tumoral/genética
10.
Nat Commun ; 11(1): 4840, 2020 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-32973129

RESUMO

Immunotherapies revolutionized cancer treatment by harnessing the immune system to target cancer cells. However, most patients are resistant to immunotherapies and the mechanisms underlying this resistant is still poorly understood. Here, we report that overexpression of BMP7, a member of the TGFB superfamily, represents a mechanism for resistance to anti-PD1 therapy in preclinical models and in patients with disease progression while on immunotherapies. BMP7 secreted by tumor cells acts on macrophages and CD4+ T cells in the tumor microenvironment, inhibiting MAPK14 expression and impairing pro-inflammatory responses. Knockdown of BMP7 or its neutralization via follistatin in combination with anti-PD1 re-sensitizes resistant tumors to immunotherapies. Thus, we identify the BMP7 signaling pathway as a potential immunotherapeutic target in cancer.


Assuntos
Proteína Morfogenética Óssea 7/genética , Proteína Morfogenética Óssea 7/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Imunoterapia/métodos , Neoplasias/metabolismo , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Linfócitos T CD4-Positivos , Linhagem Celular Tumoral , Feminino , Folistatina/metabolismo , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Proteína Quinase 14 Ativada por Mitógeno/genética , Proteína Quinase 14 Ativada por Mitógeno/metabolismo , Receptor de Morte Celular Programada 1/efeitos dos fármacos , Células RAW 264.7 , Proteína Smad1/metabolismo , Transcriptoma , Microambiente Tumoral/efeitos dos fármacos
11.
Hum Mol Genet ; 29(15): 2496-2507, 2020 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-32601694

RESUMO

In myotonic dystrophy type 1 (DM1), somatic mosaicism of the (CTG)n repeat expansion is age-dependent, tissue-specific and expansion-biased. These features contribute toward variation in disease severity and confound genotype-to-phenotype analyses. To investigate how the (CTG)n repeat expansion changes over time, we collected three longitudinal blood DNA samples separated by 8-15 years and used small pool and single-molecule PCR in 43 DM1 patients. We used the lower boundary of the allele length distribution as the best estimate for the inherited progenitor allele length (ePAL), which is itself the best predictor of disease severity. Although in most patients the lower boundary of the allele length distribution was conserved over time, in many this estimate also increased with age, suggesting samples for research studies and clinical trials should be obtained as early as possible. As expected, the modal allele length increased over time, driven primarily by ePAL, age-at-sampling and the time interval. As expected, small expansions <100 repeats did not expand as rapidly as larger alleles. However, the rate of expansion of very large alleles was not obviously proportionally higher. This may, at least in part, be a result of the allele length-dependent increase in large contractions that we also observed. We also determined that individual-specific variation in the increase of modal allele length over time not accounted for by ePAL, age-at-sampling and time was inversely associated with individual-specific variation in age-at-onset not accounted for by ePAL, further highlighting somatic expansion as a therapeutic target in DM1.


Assuntos
DNA/genética , Mosaicismo , Distrofia Miotônica/genética , Repetições de Trinucleotídeos/genética , Adolescente , Fatores Etários , Idade de Início , Alelos , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Distrofia Miotônica/patologia , Fenótipo , Expansão das Repetições de Trinucleotídeos
12.
Nucleic Acids Res ; 47(1): 152-167, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30321403

RESUMO

DNMT3L (DNMT3-like), a member of the DNMT3 family, has no DNA methyltransferase activity but regulates de novo DNA methylation. While biochemical studies show that DNMT3L is capable of interacting with both DNMT3A and DNMT3B and stimulating their enzymatic activities, genetic evidence suggests that DNMT3L is essential for DNMT3A-mediated de novo methylation in germ cells but is dispensable for de novo methylation during embryogenesis, which is mainly mediated by DNMT3B. How DNMT3L regulates DNA methylation and what determines its functional specificity are not well understood. Here we show that DNMT3L-deficient mouse embryonic stem cells (mESCs) exhibit downregulation of DNMT3A, especially DNMT3A2, the predominant DNMT3A isoform in mESCs. DNA methylation analysis of DNMT3L-deficient mESCs reveals hypomethylation at many DNMT3A target regions. These results confirm that DNMT3L is a positive regulator of DNA methylation, contrary to a previous report that, in mESCs, DNMT3L regulates DNA methylation positively or negatively, depending on genomic regions. Mechanistically, DNMT3L forms a complex with DNMT3A2 and prevents DNMT3A2 from being degraded. Restoring the DNMT3A protein level in DNMT3L-deficient mESCs partially recovers DNA methylation. Thus, our work uncovers a role for DNMT3L in maintaining DNMT3A stability, which contributes to the effect of DNMT3L on DNMT3A-dependent DNA methylation.


Assuntos
DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA/genética , Desenvolvimento Embrionário/genética , Animais , DNA Metiltransferase 3A , Estabilidade Enzimática/genética , Camundongos , Células-Tronco Embrionárias Murinas/metabolismo , Ligação Proteica/genética
13.
Cancer Res ; 77(9): 2500-2511, 2017 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-28373182

RESUMO

The association between obesity and breast cancer risk and prognosis is well established in estrogen receptor (ER)-positive disease but less clear in HER2-positive disease. Here, we report preclinical evidence suggesting weight maintenance through calorie restriction (CR) may limit risk of HER2-positive breast cancer. In female MMTV-HER2/neu transgenic mice, we found that ERα and ERß expression, mammary tumorigenesis, and survival are energy balance dependent in association with epigenetic reprogramming. Mice were randomized to receive a CR, overweight-inducing, or diet-induced obesity regimen (n = 27/group). Subsets of mice (n = 4/group/time point) were euthanized after 1, 3, and 5 months to characterize diet-dependent metabolic, transcriptional, and epigenetic perturbations. Remaining mice were followed up to 22 months. Relative to the overweight and diet-induced obesity regimens, CR decreased body weight, adiposity, and serum metabolic hormones as expected and also elicited an increase in mammary ERα and ERß expression. Increased DNA methylation accompanied this pattern, particularly at CpG dinucleotides located within binding or flanking regions for the transcriptional regulator CCCTC-binding factor of ESR1 and ESR2, consistent with sustained transcriptional activation of ERα and ERß. Mammary expression of the DNA methylation enzyme DNMT1 was stable in CR mice but increased over time in overweight and diet-induced obesity mice, suggesting CR obviates epigenetic alterations concurrent with chronic excess energy intake. In the survival study, CR elicited a significant suppression in spontaneous mammary tumorigenesis. Overall, our findings suggest a mechanistic rationale to prevent or reverse excess body weight as a strategy to reduce HER2-positive breast cancer risk. Cancer Res; 77(9); 2500-11. ©2017 AACR.


Assuntos
Neoplasias da Mama/genética , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Neoplasias Mamárias Animais/genética , Obesidade/genética , Animais , Neoplasias da Mama/fisiopatologia , Restrição Calórica , Carcinogênese/genética , Metilação de DNA/genética , Metabolismo Energético/genética , Epigênese Genética/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Mamárias Animais/etiologia , Neoplasias Mamárias Animais/fisiopatologia , Camundongos , Camundongos Transgênicos , Obesidade/complicações , Obesidade/fisiopatologia , Receptor ErbB-2/genética , Fatores de Risco
14.
Epigenetics ; 11(3): 184-93, 2016 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-26890396

RESUMO

Small cell prostate carcinoma (SCPC) morphology is rare at initial diagnosis but often emerges during prostate cancer progression and portends a dismal prognosis. It does not express androgen receptor (AR) or respond to hormonal therapies. Clinically applicable markers for its early detection and treatment with effective chemotherapy are needed. Our studies in patient tumor-derived xenografts (PDX) revealed that AR-negative SCPC (AR(-)SCPC) expresses neural development genes instead of the prostate luminal epithelial genes characteristic of AR-positive castration-resistant adenocarcinomas (AR(+)ADENO). We hypothesized that the differences in cellular lineage programs are reflected in distinct epigenetic profiles. To address this hypothesis, we compared the DNA methylation profiles of AR(-) and AR(+) PDX using methylated CpG island amplification and microarray (MCAM) analysis and identified a set of differentially methylated promoters, validated in PDX and corresponding donor patient samples. We used the Illumina 450K platform to examine additional regions of the genome and the correlation between the DNA methylation profiles of the PDX and their corresponding patient tumors. Struck by the low frequency of AR promoter methylation in the AR(-)SCPC, we investigated this region's specific histone modification patterns by chromatin immunoprecipitation. We found that the AR promoter was enriched in silencing histone modifications (H3K27me3 and H3K9me2) and that EZH2 inhibition with 3-deazaneplanocin A (DZNep) resulted in AR expression and growth inhibition in AR(-)SCPC cell lines. We conclude that the epigenome of AR(-) is distinct from that of AR(+) castration-resistant prostate carcinomas, and that the AR(-) phenotype can be reversed with epigenetic drugs.


Assuntos
Carcinoma de Células Pequenas/genética , Metilação de DNA/genética , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/genética , Adenosina/administração & dosagem , Adenosina/análogos & derivados , Animais , Carcinoma de Células Pequenas/patologia , Linhagem Celular Tumoral , Linhagem da Célula/genética , Ilhas de CpG/genética , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Camundongos , Regiões Promotoras Genéticas , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/biossíntese , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Cancer Prev Res (Phila) ; 9(5): 339-48, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-26869351

RESUMO

Using a murine model of basal-like breast cancer, we tested the hypothesis that chronic obesity, an established breast cancer risk and progression factor in women, induces mammary gland epigenetic reprogramming and increases mammary tumor growth. Moreover, we assessed whether the obesity-induced epigenetic and protumor effects are reversed by weight normalization. Ovariectomized female C57BL/6 mice were fed a control diet or diet-induced obesity (DIO) regimen for 17 weeks, resulting in a normal weight or obese phenotype, respectively. Mice on the DIO regimen were then randomized to continue the DIO diet or were switched to the control diet, resulting in formerly obese (FOb) mice with weights comparable with control mice. At week 24, all mice were orthotopically injected with MMTV-Wnt-1 mouse mammary tumor cells. Mean tumor volume, serum IL6 levels, expression of proinflammatory genes in the mammary fat pad, and mammary DNA methylation profiles were similar in DIO and FOb mice and higher than in controls. Many of the genes found to have obesity-associated hypermethylation in mice were also found to be hypermethylated in the normal breast tissue of obese versus nonobese human subjects, and nearly all of these concordant genes remained hypermethylated after significant weight loss in the FOb mice. Our findings suggest that weight normalization may not be sufficient to reverse the effects of chronic obesity on epigenetic reprogramming and inflammatory signals in the microenvironment that are associated with breast cancer progression. Cancer Prev Res; 9(5); 339-48. ©2016 AACR.


Assuntos
Epigênese Genética , Neoplasias Mamárias Experimentais/etiologia , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Obesidade/complicações , Animais , Carcinoma Ductal de Mama/genética , Carcinoma Ductal de Mama/patologia , Metilação de DNA/genética , Modelos Animais de Doenças , Feminino , Humanos , Inflamação/complicações , Camundongos , Camundongos Endogâmicos C57BL , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase
16.
J Neurosci ; 35(34): 12002-17, 2015 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-26311780

RESUMO

Lamin B1 is a component of the nuclear lamina and plays a critical role in maintaining nuclear architecture, regulating gene expression and modulating chromatin positioning. We have previously shown that LMNB1 gene duplications cause autosomal dominant leukodystrophy (ADLD), a fatal adult onset demyelinating disease. The mechanisms by which increased LMNB1 levels cause ADLD are unclear. To address this, we used a transgenic mouse model where Lamin B1 overexpression is targeted to oligodendrocytes. These mice showed severe vacuolar degeneration of the spinal cord white matter together with marked astrogliosis, microglial infiltration, and secondary axonal damage. Oligodendrocytes in the transgenic mice revealed alterations in histone modifications favoring a transcriptionally repressed state. Chromatin changes were accompanied by reduced expression of genes involved in lipid synthesis pathways, many of which are known to play important roles in myelin regulation and are preferentially expressed in oligodendrocytes. Decreased lipogenic gene expression resulted in a significant reduction in multiple classes of lipids involved in myelin formation. Many of these gene expression changes and lipid alterations were observed even before the onset of the phenotype, suggesting a causal role. Our findings establish, for the first time, a link between LMNB1 and lipid synthesis in oligodendrocytes, and provide a mechanistic framework to explain the age dependence and white matter involvement of the disease phenotype. These results have implications for disease pathogenesis and may also shed light on the regulation of lipid synthesis pathways in myelin maintenance and turnover. SIGNIFICANCE STATEMENT: Autosomal dominant leukodystrophy (ADLD) is fatal neurological disorder caused by increased levels of the nuclear protein, Lamin B1. The disease is characterized by an age-dependent loss of myelin, the fatty sheath that covers nerve fibers. We have studied a mouse model where Lamin B1 level are increased in oligodendrocytes, the cell type that produces myelin in the CNS. We demonstrate that destruction of myelin in the spinal cord is responsible for the degenerative phenotype in our mouse model. We show that this degeneration is mediated by reduced expression of lipid synthesis genes and the subsequent reduction in myelin enriched lipids. These findings provide a mechanistic framework to explain the age dependence and tissue specificity of the ADLD disease phenotype.


Assuntos
Envelhecimento/metabolismo , Doenças Desmielinizantes/metabolismo , Lamina Tipo B/biossíntese , Metabolismo dos Lipídeos/fisiologia , Envelhecimento/genética , Animais , Doenças Desmielinizantes/genética , Regulação da Expressão Gênica , Humanos , Camundongos , Camundongos Transgênicos , Lâmina Nuclear/genética , Lâmina Nuclear/metabolismo , Oligodendroglia/metabolismo
17.
Cancer Res ; 75(18): 3980-90, 2015 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-26249178

RESUMO

Ductal carcinoma in situ (DCIS) is a noninvasive precursor lesion to invasive breast carcinoma. We still have no understanding on why only some DCIS lesions evolve to invasive cancer whereas others appear not to do so during the life span of the patient. Here, we performed full exome (tumor vs. matching normal), transcriptome, and methylome analysis of 30 pure high-grade DCIS (HG-DCIS) and 10 normal breast epithelial samples. Sixty-two percent of HG-DCIS cases displayed mutations affecting cancer driver genes or potential drivers. Mutations were observed affecting PIK3CA (21% of cases), TP53 (17%), GATA3 (7%), MLL3 (7%) and single cases of mutations affecting CDH1, MAP2K4, TBX3, NF1, ATM, and ARID1A. Significantly, 83% of lesions displayed numerous large chromosomal copy number alterations, suggesting they might precede selection of cancer driver mutations. Integrated pathway-based modeling analysis of RNA-seq data allowed us to identify two DCIS subgroups (DCIS-C1 and DCIS-C2) based on their tumor-intrinsic subtypes, proliferative, immune scores, and in the activity of specific signaling pathways. The more aggressive DCIS-C1 (highly proliferative, basal-like, or ERBB2(+)) displayed signatures characteristic of activated Treg cells (CD4(+)/CD25(+)/FOXP3(+)) and CTLA4(+)/CD86(+) complexes indicative of a tumor-associated immunosuppressive phenotype. Strikingly, all lesions showed evidence of TP53 pathway inactivation. Similarly, ncRNA and methylation profiles reproduce changes observed postinvasion. Among the most significant findings, we observed upregulation of lncRNA HOTAIR in DCIS-C1 lesions and hypermethylation of HOXA5 and SOX genes. We conclude that most HG-DCIS lesions, in spite of representing a preinvasive stage of tumor progression, displayed molecular profiles indistinguishable from invasive breast cancer.


Assuntos
Neoplasias da Mama/genética , Carcinoma Intraductal não Infiltrante/química , Metilação de DNA , DNA de Neoplasias/genética , Perfilação da Expressão Gênica , Proteínas de Neoplasias/genética , Transcriptoma , Antígenos de Diferenciação de Linfócitos T/análise , Mama/química , Neoplasias da Mama/química , Neoplasias da Mama/imunologia , Antígeno CTLA-4/análise , Carcinoma Intraductal não Infiltrante/classificação , Carcinoma Intraductal não Infiltrante/genética , Carcinoma Intraductal não Infiltrante/imunologia , Feminino , Regulação Neoplásica da Expressão Gênica , Genes Neoplásicos , Humanos , Linfócitos do Interstício Tumoral/química , Linfócitos do Interstício Tumoral/imunologia , Mutação , Invasividade Neoplásica/genética , Proteínas de Neoplasias/análise , RNA Mensageiro/genética , RNA Neoplásico/genética , RNA não Traduzido/genética , Linfócitos T Reguladores/imunologia
18.
Cancer Res ; 75(14): 2833-43, 2015 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-25972343

RESUMO

TET2 enzymatically converts 5-methylcytosine to 5-hydroxymethylcytosine as well as other covalently modified cytosines and its mutations are common in myeloid leukemia. However, the exact mechanism and the extent to which TET2 mutations affect DNA methylation remain in question. Here, we report on DNA methylomes in TET2 wild-type (TET2-WT) and mutant (TET2-MT) cases of chronic myelomonocytic leukemia (CMML). We analyzed 85,134 CpG sites [28,114 sites in CpG islands (CGI) and 57,020 in non-CpG islands (NCGI)]. TET2 mutations do not explain genome-wide differences in DNA methylation in CMML, and we found few and inconsistent differences at CGIs between TET2-WT and TET2-MT cases. In contrast, we identified 409 (0.71%) TET2-specific differentially methylated CpGs (tet2-DMCs) in NCGIs, 86% of which were hypermethylated in TET2-MT cases, suggesting a strikingly different biology of the effects of TET2 mutations at CGIs and NCGIs. DNA methylation of tet2-DMCs at promoters and nonpromoters repressed gene expression. Tet2-DMCs showed significant enrichment at hematopoietic-specific enhancers marked by H3K4me1 and at binding sites for the transcription factor p300. Tet2-DMCs showed significantly lower 5-hydroxymethylcytosine in TET2-MT cases. We conclude that leukemia-associated TET2 mutations affect DNA methylation at NCGI regions containing hematopoietic-specific enhancers and transcription factor-binding sites.


Assuntos
Metilação de DNA/genética , Proteínas de Ligação a DNA/genética , Elementos Facilitadores Genéticos/genética , Leucemia Mielomonocítica Crônica/genética , Mutação de Sentido Incorreto , Proteínas Proto-Oncogênicas/genética , Fatores de Transcrição/metabolismo , Sítios de Ligação/genética , Estudos de Casos e Controles , Ilhas de CpG , Dioxigenases , Perfilação da Expressão Gênica , Regulação Leucêmica da Expressão Gênica , Genoma Humano , Hematopoese/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Análise em Microsséries , Regiões Promotoras Genéticas/genética
19.
Tumour Biol ; 36(10): 8147-58, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25990457

RESUMO

AURKC, a member of the Aurora kinase gene family, is highly expressed in testis but is either moderately expressed or repressed in most somatic cells. Varying expression of AURKC has been observed in human cancers, but the underlying mechanisms of differential expression have been investigated only to a limited extent. We investigated the role of promoter CpG methylation in the regulation of AURKC gene expression in human cancer cells, in relation to a recently reported AURKC transcription repressor PLZF/ZBTB16, implicated in transformation and tumorigenesis. AURKC and PLZF/ZBTB16 expression profiles were investigated in reference to CpG methylation status on the AURKC promoter experimentally, and also in The Cancer Genome Atlas (TCGA) dataset involving multiple cancer types. AURKC promoter showed dense to moderate hypermethylation correlating with low to moderate expression of the gene in normal somatic cells and cancer cell lines, while testis with high expression revealed marked hypo-methylation. Treatment with the demethylating agent, 5-aza-dC, but not the histone deacetylase (HDAC) inhibitor, TSA, led to elevated expression in cancer cell lines, indicating that promoter DNA methylation negatively regulates AURKC expression. High expression of PLZF in PLZF-transfected cells treated with 5-aza-dC only partially repressed expression of AURKC despite 5-aza-dC also inducing elevated PLZF expression. Analyses of the TCGA data showed differential expression of AURKC in multiple cancer types and stronger correlation of AURKC expression with CpG methylation compared to PLZF levels. These findings demonstrate that differential promoter CpG methylation is an important mechanism regulating AURKC expression in cancer cells.


Assuntos
Aurora Quinase C/genética , Ilhas de CpG/genética , Metilação de DNA , Regulação Neoplásica da Expressão Gênica , Neoplasias/genética , Testículo/metabolismo , Transformação Celular Neoplásica , Humanos , Masculino , Neoplasias/patologia , Regiões Promotoras Genéticas/genética , Reação em Cadeia da Polimerase em Tempo Real
20.
Contemp Oncol (Pozn) ; 19(1A): A30-8, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25691819

RESUMO

Reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) requires profound alterations in the epigenetic landscape. During reprogramming, a change in chromatin structure resets the gene expression and stabilises self-renewal. Reprogramming is a highly inefficient process, in part due to multiple epigenetic barriers. Although many epigenetic factors have already been shown to affect self-renewal and pluripotency in embryonic stem cells (ESCs), only a few of them have been examined in the context of dedifferentiation. In order to improve current protocols of iPSCs generation, it is essential to identify epigenetic drivers and blockages of somatic cell reprogramming.

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