RESUMO
Cell-free DNA (cfDNA) analysis, specifically circulating tumor DNA (ctDNA) analysis, provides enormous opportunities for noninvasive early assessment of cancers. To date, PCR-based methods have led this field. However, the limited sensitivity/specificity of PCR-based methods necessitates the search for new methods. Here, we describe a direct approach to detect KRAS G12D mutated genes in clinical ctDNA samples with the utmost LOD and sensitivity/specificity. In this study, MutS protein was immobilized on the tip of an atomic force microscope (AFM), and the protein sensed the mismatched sites of the duplex formed between the capture probe on the surface and mutated DNA. A noteworthy LOD (3 copies, 0.006% allele frequency) was achieved, along with superb sensitivity/specificity (100%/100%). These observations demonstrate that force-based AFM, in combination with the protein found in nature and properly designed capture probes/blockers, represents an exciting new avenue for ctDNA analysis.
Assuntos
DNA Tumoral Circulante , Neoplasias , Biomarcadores Tumorais/genética , DNA Tumoral Circulante/genética , Humanos , Mutação , Mutação Puntual , Sensibilidade e EspecificidadeRESUMO
Tumor heterogeneity is one of the ongoing huddles in the field of colon cancer therapy. It is evident that there are countless clones which exhibit different phenotypes and therefore, single cell analysis is inevitable. Cancer stem cells (CSCs) are rare cell population within tumor which is known to function in cancer metastasis and recurrence. Although there have been trials to prove intra-tumoral heterogeneity using single cell sequencing, that of CSCs has not been clearly elucidated. Here, we articulate the presence of heterogeneous subclones within CD133 positive cancer stem cells through single cell sequencing. As a proof of principle, we performed phenotype-based high-throughput laser isolation and single cell sequencing (PHLI-seq) of CD133 positive cells in a frozen tumor tissue obtained from a patient with colorectal cancer. The result proved that CD133 positive cells were shown to be heterogeneous both in copy number and mutational profiles. Single cancer stem cell specific mutations such as RNF144A, PAK2, PARP4, ADAM21, HYDIN, KRT38 and CELSR1 could be also detected in liver metastatic tumor of the same patient. Collectively, these data suggest that single cell analysis used to spot subclones with genetic variation within rare population, will lead to new strategies to tackle colon cancer metastasis.
Assuntos
Antígeno AC133/metabolismo , Células-Tronco Neoplásicas/classificação , Células-Tronco Neoplásicas/metabolismo , Idoso , Biomarcadores Tumorais/metabolismo , Separação Celular/métodos , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Dosagem de Genes , Humanos , Lasers , Masculino , Mutação , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Células-Tronco Neoplásicas/patologia , Fenótipo , Análise de Célula Única , Sequenciamento do ExomaRESUMO
BACKGROUND: Gastric cancer (GC) is a leading cause of cancer morbidity and mortality worldwide. This is due to the heterogeneous features of GC, which consist of a diverse molecular phenotype. Epstein-Barr virus (EBV)-positive GC and microsatellite instability (MSI)-high GC encompass similar epigenetic traits, including high levels of DNA methylation in CpG islands; however, EBV-positive and MSI-high GCs are mutually exclusive. We aimed to elucidate the underlying mechanism of this exclusivity. METHODS: We knocked out MLH1 in EBV-positive GC cell lines SNU-719 and NCC24 via CRISPR-Cas9, and evaluated the modified cellular properties in vitro and in vivo. The MSI status of each cell line was screened with two marker capillary electrophoresis, and further diagnosed with five marker capillary electrophoresis and parallel sequencing using 21 markers. RESULTS: Initial evaluation showed that cell growth, migration, invasion, and MSI status were not affected by MLH1 silencing. However, with prolonged passage, GC cell lines gradually gained MSI and NCC24 cells were transformed to EBV-positive/MSI-high GC cells after 12 months. Furthermore, MLH1 silencing reduced tumor stemness in SNU-719 and NCC24 regardless of the MSI status in vitro and in vivo. CONCLUSIONS: Our findings suggest that EBV-positivity and MSI-high status are mutually exclusive due to the immediate disadvantage in tumor stemness when MLH1 is silenced, whereas the establishment of MSI-high status in EBV-positive GCs required a longer period.
Assuntos
Herpesvirus Humano 4/isolamento & purificação , Instabilidade de Microssatélites , Neoplasias Gástricas/patologia , Transformação Celular Neoplásica , Humanos , Neoplasias Gástricas/virologiaRESUMO
The epithelial-mesenchymal transition (EMT) comprises an important biological mechanism not only for cancer progression but also in the therapeutic resistance of cancer cells. While the importance of the protein abundance of EMT-inducers, such as Snail (SNAI1) and Zeb1 (ZEB1), during EMT progression is clear, the reciprocal interactions between the untranslated regions (UTRs) of EMT-inducers via a competing endogenous RNA (ceRNA) network have received little attention. In this study, we found a synchronized transcript abundance of Snail and Zeb1 mediated by a non-coding RNA network in colorectal cancer (CRC). Importantly, the trans-regulatory ceRNA network in the UTRs of EMT inducers is mediated by competition between tumor suppressive miRNA-34 (miR-34) and miRNA-200 (miR-200). Furthermore, the ceRNA network consisting of the UTRs of EMT inducers and tumor suppressive miRs is functional in the EMT phenotype and therapeutic resistance of colon cancer. In The Cancer Genome Atlas (TCGA) samples, we also found genome-wide ceRNA gene sets regulated by miR-34a and miR-200 in colorectal cancer. These results indicate that the ceRNA networks regulated by the reciprocal interaction between EMT gene UTRs and tumor suppressive miRs are functional in CRC progression and therapeutic resistance.
Assuntos
Neoplasias Colorretais/metabolismo , Genes Supressores de Tumor , MicroRNAs/metabolismo , Proteínas de Neoplasias/metabolismo , RNA Neoplásico/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Animais , Neoplasias Colorretais/genética , Neoplasias Colorretais/terapia , Feminino , Células HCT116 , Humanos , Camundongos , Camundongos Nus , MicroRNAs/genética , Proteínas de Neoplasias/genética , RNA Neoplásico/genética , Fatores de Transcrição da Família Snail/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genéticaRESUMO
PIK3CA is a frequently mutated gene in cancer, including about ~15 to 20% of colorectal cancers (CRC). PIK3CA mutations lead to activation of the PI3K/AKT/mTOR signaling pathway, which plays pivotal roles in tumorigenesis. Here, we investigated the mechanism of resistance of PIK3CA-mutant CRC cell lines to gedatolisib, a dual PI3K/mTOR inhibitor. Out of a panel of 29 CRC cell lines, we identified 7 harboring one or more PIK3CA mutations; of these, 5 and 2 were found to be sensitive and resistant to gedatolisib, respectively. Both of the gedatolisib-resistant cell lines expressed high levels of active glycogen synthase kinase 3-beta (GSK3ß) and harbored the same frameshift mutation (c.465_466insC; H155fs*) in TCF7, which encodes a positive transcriptional regulator of the WNT/ß-catenin signaling pathway. Inhibition of GSK3ß activity in gedatolisib-resistant cells by siRNA-mediated knockdown or treatment with a GSK3ß-specific inhibitor effectively reduced the activity of molecules downstream of mTOR and also decreased signaling through the WNT/ß-catenin pathway. Notably, GSK3ß inhibition rendered the resistant cell lines sensitive to gedatolisib cytotoxicity, both in vitro and in a mouse xenograft model. Taken together, these data demonstrate that aberrant regulation of WNT/ß-catenin signaling and active GSK3ß induced by the TCF7 frameshift mutation cause resistance to the dual PI3K/mTOR inhibitor gedatolisib. Cotreatment with GSK3ß inhibitors may be a strategy to overcome the resistance of PIK3CA- and TCF7-mutant CRC to PI3K/mTOR-targeted therapies.
Assuntos
Classe I de Fosfatidilinositol 3-Quinases/genética , Neoplasias Colorretais/metabolismo , Resistencia a Medicamentos Antineoplásicos/fisiologia , Morfolinas/farmacologia , Triazinas/farmacologia , Via de Sinalização Wnt/fisiologia , Animais , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Humanos , Camundongos , Mutação , Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Some clinicians are concerned that if an intraoral vertical ramus osteotomy (IVRO) is used to position the mandible posteriorly, the proximal segments should be positioned laterally to the distal segment, which could increase the transverse mandibular width, leading to esthetically unfavorable results. This study investigated short- and long-term postoperative transverse mandibular width changes in the soft and hard tissue after IVRO for mandibular prognathism. MATERIALS AND METHODS: The study comprised 44 patients who were treated with mandibular setback surgery using an IVRO. They were categorized into either the facial symmetry group or facial asymmetry group based on their preoperative levels of chin top deviation. Three-dimensional cone-beam computed tomography images were obtained at the preoperative, 1-month postoperative, and 12-month postoperative stages, designated as T1, T2, and T3, respectively. We set hard tissue width 1 (HW1) and hard tissue width 2 (HW2) as the sum of the distance at the bilateral ends of the angle and ramus, respectively, and set soft tissue width 1 (SW1) and soft tissue width 2 (SW2) as the sum of the distance at the bilateral ends of the soft tissue angle and ramus, respectively. RESULTS: Compared with the value at T1, the HW1 value increased by 8.16% (P < .05) and HW2 increased by 4.39% (P > .05) at T2; HW1 increased by 4.35% (P < .05) and HW2 increased by 2.95% (P > .05) at T3. Compared with the value at T1, the SW1 value increased by 2.49% and SW2 increased by 2.50% at T2; however, SW1 decreased by 0.85% and SW2 increased by 0.37% at T3. The soft tissue variations between T1 and T2, as well as between T2 and T3, were statistically significant. However, no significant difference was found between T1 and T3 (P > .05). No difference between the facially symmetrical and asymmetrical groups was found over time for soft and hard tissues (P > .05). CONCLUSIONS: Notably, IVRO does not seem to impact the transverse facial profile and enables reliable prediction of the esthetic results of surgery.
Assuntos
Mandíbula/cirurgia , Osteotomia Sagital do Ramo Mandibular , Cefalometria , Estética Dentária , Seguimentos , Humanos , Osteotomia Mandibular , Prognatismo , Estudos RetrospectivosRESUMO
Epithelial to mesenchymal transition (EMT) and mesenchymal to epithelial transition (MET) are important interconnected events in tumorigenesis controlled by complex genetic networks. However, the cues that activate EMT-initiating factors and the mechanisms that reversibly connect EMT/MET are not well understood. Here, we show that cohesin-mediated chromatin organization coordinates EMT/MET by regulating mesenchymal genes. We report that RAD21, a subunit of the cohesin complex, is expressed in epithelial breast cancer cells, whereas its expression is decreased in mesenchymal cancer. Depletion of RAD21 in epithelial cancer cells causes transcriptional activation of TGFB1 and ITGA5, inducing EMT. Reduced binding of RAD21 changes intrachromosomal chromatin interactions within the TGFB1 and ITGA5 loci, creating an active transcriptional environment. Similarly, stem cell-like cancer cells also show an open chromatin structure at both genes, which correlates with high expression levels and mesenchymal fate characteristics. Conversely, overexpression of RAD21 in mesenchymal cancer cells induces MET-specific expression patterns. These findings indicate that dynamic cohesin-mediated chromatin structures are responsible for the initiation and regulation of essential EMT-related cell fate changes in cancer.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Cromatina/genética , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Transição Epitelial-Mesenquimal/genética , Neoplasias/genética , Neoplasias/metabolismo , Linhagem Celular Tumoral , Montagem e Desmontagem da Cromatina/genética , Proteínas de Ligação a DNA , Células Epiteliais/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Metástase Neoplásica , Neoplasias/patologia , Proteínas Nucleares/genética , Fosfoproteínas/genética , Regiões Promotoras Genéticas , Fator de Crescimento Transformador beta1/genética , CoesinasRESUMO
Gene amplification is a hallmark of cancer with chromosomal instability although the underlying mechanism by which altered copy numbers are maintained is largely unclear. Cohesin, involved in sister chromatid cohesion, DNA repair, cell cycle progression and transcriptional regulation of key developmental genes, is frequently overexpressed in human cancer. Here we show that cohesin-dependent change in DNA replication controls the copy numbers of amplified genes in cancer cells with chromosomal instability. We found that the down-regulation of elevated cohesin leads to copy number-associated gene expression changes without disturbing chromosomal segregation. Highly amplified genes form typical long-range chromatin interactions, which are stabilized by enriched cohesin. The spatial proximities among cohesin binding sites within amplified genes are decreased by RAD21-knockdown, resulting in the rapid decline of amplified gene expression. After several passages, cohesin depletion inhibits DNA replication initiation by reducing the recruitment of pre-replication complexes such as minichromosome maintenance subunits 7 (MCM7), DNA polymerase α, and CDC45 at replication origins near the amplified regions, and as a result, decreases the DNA copy numbers of highly amplified genes. Collectively, our data demonstrate that cohesin-mediated chromatin organization and DNA replication are important for stabilizing gene amplification in cancer cells with chromosomal instability.
Assuntos
Instabilidade Cromossômica , Amplificação de Genes , Regulação Neoplásica da Expressão Gênica , Proteínas Nucleares/genética , Fosfoproteínas/genética , Neoplasias Gástricas/genética , Sítios de Ligação , Fator de Ligação a CCCTC , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Cromátides/química , Cromátides/metabolismo , Cromatina/química , Cromatina/metabolismo , Segregação de Cromossomos , Hibridização Genômica Comparativa , DNA Polimerase I/genética , DNA Polimerase I/metabolismo , Replicação do DNA , Proteínas de Ligação a DNA , Dosagem de Genes , Células HCT116 , Células Hep G2 , Humanos , Componente 7 do Complexo de Manutenção de Minicromossomo/genética , Componente 7 do Complexo de Manutenção de Minicromossomo/metabolismo , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/metabolismo , Fosfoproteínas/antagonistas & inibidores , Fosfoproteínas/metabolismo , Ligação Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologiaRESUMO
Recent genome-wide epigenomic and transcription profiling studies have demonstrated that epigenetic silencing can encompass multiple neighboring genes, termed as long-range epigenetic silencing (LRES). Herein, we identified a novel LRES region by comparing gene expression of human colon cancer HCT116 cells with their DNA methyltransferase 1 and DNA methyltransferase 3B double-knockout derivative double-knockout cells. Ten consecutive genes spanning 3 Mb of chromosome 15q25 were coordinately silenced, with eight genes showing promoter CpG island hypermethylation and enrichment of repressive histone marks, which were evaluated by bisulfite sequencing analysis and chromatin immunoprecipitation assay. Comparison of primary gastric tumor specimens with normal tissue confirmed that the long-range silencing of this region was tumor specific. Methylation of genes within the LRES region was evaluated in 190 gastric tumor tissues using the MethyLight assay, and their association with clinicopathological features, such as older age, high-grade differentiation, and diffuse or mixed-type histology, was determined. LRES-positive gastric cancer patients (six or more methylated genes) showed lower recurrence and better survival. Our findings emphasize the differential dynamics of DNA methylation and histone modification, indicating the importance of studying the relationship of each epigenetic modification in the context of chromatin domains. Patients with LRES showed lower recurrence and better prognosis, indicating that stratifying patients according to underlying molecular features, such as LRES regions, may better predict recurrence and survival.
Assuntos
Cromossomos Humanos Par 15 , Epigênese Genética , Inativação Gênica , Recidiva Local de Neoplasia/genética , Neoplasias Gástricas/genética , Adulto , Idoso , Ilhas de CpG , Metilação de DNA , Feminino , Histonas , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/mortalidade , Recidiva Local de Neoplasia/patologia , Regiões Promotoras Genéticas , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Taxa de Sobrevida , Adulto JovemRESUMO
INTRODUCTION: Olaparib, a poly (ADP-ribose) polymerase (PARP) inhibitor, has been found to have therapeutic potential for treating cancers associated with impaired DNA repair capabilities, particularly those with deficiencies in the homologous recombination repair (HRR) pathway. Histone deacetylases (HDACs) are important for enabling functional HRR of DNA by regulating the expression of HRR-related genes and promoting the accurate assembly of HRR-directed sub-nuclear foci. Thus, HDAC inhibitors have recently emerged as a therapeutic agent for treating cancer by inhibiting DNA repair. Based on this, HDAC inhibition could be predicted to enhance the anti-tumor effect of PARP inhibitors in cancer cells by blocking the HRR pathway. METHODS: We determined whether suberoylanilide hydroxamic acid (SAHA), a HDAC inhibitor, could enhance the anti-tumor effects of olaparib on breast cancer cell lines using a cytotoxic assay, cell cycle analysis, and Western blotting. We evaluated how exposure to SAHA affects the expression of HRR-associated genes. The accumulation of DNA double strand breaks (DSBs) induced by combination treatment was assessed. Induction of autophagy was monitored by imaging green fluorescent protein-tagged microtubule-associated protein 1A/1B-light chain 3 (LC3) expression following co-treatment with olaparib and SAHA. These in vitro data were validated in vivo using a human breast cancer xenograft model. RESULTS: Triple-negative breast cancer cell (TNBC) lines showed heterogeneous responses to the PARP and HDAC inhibitors. Co-administration of olaparib and SAHA synergistically inhibited the growth of TNBC cells that expressed functional Phosphatase and tensin homolog (PTEN). This effect was associated with down-regulation of the proliferative signaling pathway, increased apoptotic and autophagic cell death, and accumulation of DNA damage. The combined anti-tumor effect of olaparib and SAHA was also observed in a xenograft model. These data suggest that PTEN expression in TNBC cells can sensitize the cell response to simultaneous inhibition of PARP and HDAC both in vitro and in vivo. CONCLUSION: Our findings suggest that expression of functional PTEN may serve as a biomarker for selecting TNBC patients that would favorably respond to a combination of olaparib with SAHA. This provides a strong rationale for treating TNBC patients with PTEN expression with a combination therapy consisting of olaparib and SAHA.
Assuntos
Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Ftalazinas/farmacologia , Piperazinas/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Animais , Apoptose , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/genética , Sinergismo Farmacológico , Feminino , Expressão Gênica , Humanos , Camundongos , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Vorinostat , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Erythropoiesis is dependent on the lineage-specific transcription factors Gata1, Tal1, and Klf1. Several erythroid genes have been shown to require all 3 factors for their expression, suggesting that they function synergistically; however, there is little direct evidence for widespread cooperation. Gata1 and Tal1 can assemble within higher-order protein complexes (Ldb1 complexes) that include the adapter molecules Lmo2 and Ldb1. Ldb1 proteins are capable of coassociation, and long-range Ldb1-mediated oligomerization of enhancer- and promoter-bound Ldb1 complexes has been shown to be required for ß-globin gene expression. In this study, we generated a genomewide map of Ldb1 complex binding sites that revealed widespread binding at erythroid genes and at known erythroid enhancer elements. Ldb1 complex binding sites frequently colocalized with Klf1 binding sites and with consensus binding motifs for other erythroid transcription factors. Transcriptomic analysis demonstrated a strong correlation between Ldb1 complex binding and Ldb1 dependency for gene expression and identified a large cohort of genes coregulated by Ldb1 complexes and Klf1. Together, these results provide a foundation for defining the mechanism and scope of Ldb1 complex activity during erythropoiesis.
Assuntos
Proteínas de Ligação a DNA/genética , Células Eritroides/metabolismo , Fator de Transcrição GATA1/genética , Proteínas com Domínio LIM/genética , Transcrição Gênica/fisiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Sítios de Ligação/genética , Células da Medula Óssea/citologia , Células da Medula Óssea/fisiologia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Células Eritroides/citologia , Eritropoese/genética , Eritropoese/fisiologia , Fator de Transcrição GATA1/metabolismo , Regulação da Expressão Gênica/fisiologia , Teste de Complementação Genética , Sequenciamento de Nucleotídeos em Larga Escala , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Proteínas com Domínio LIM/metabolismo , Leucemia Eritroblástica Aguda , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteína 1 de Leucemia Linfocítica Aguda de Células TRESUMO
Carbonic anhydrase 1 (Car1), an early specific marker of the erythroid differentiation, has been used to distinguish fetal and adult erythroid cells since its production closely follows the γ- to ß-globin transition, but the molecular mechanism underlying transcriptional regulation of Car1 is unclear. Here, we show that Car1 mRNA decreases significantly when erythroid differentiation is induced in MEL cells. The Ldb1 protein complex including GATA1/SCL/LMO2 binds to the Car1 promoter in uninduced cells and reduced enrichment of the complex during differentiation correlates with loss of Car1 expression. Knockdown of Ldb1 results in a reduction of Ser2 phosphorylated RNA Pol II and Cdk9 at the Car1 promoter region, suggesting that Ldb1 is required for recruitment of Pol II as well as the transcription regulator P-TEFb to enhance elongation of Car1 transcripts. Taken together, these data show that Ldb1 forms a regulatory complex to maintain Car1 expression in erythroid cells.
Assuntos
Anidrase Carbônica I , Proteínas de Ligação a DNA , Células Eritroides , Globinas , Proteínas com Domínio LIM , Animais , Anidrase Carbônica I/genética , Anidrase Carbônica I/metabolismo , Diferenciação Celular , Linhagem Celular , Quinase 9 Dependente de Ciclina/genética , Quinase 9 Dependente de Ciclina/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Células Eritroides/citologia , Células Eritroides/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Globinas/química , Globinas/genética , Globinas/metabolismo , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Camundongos , Fosforilação , Fator B de Elongação Transcricional Positiva/metabolismo , Regiões Promotoras GenéticasRESUMO
The Y-shaped, low molecular mass, hole-conductor (HC), acidic coadsorbents 4-{3,7-bis[4-(2-ethylhexyloxy)phenyl]-10H-phenothiazin-10-yl}benzoic acid (PTZ1) and 4-{3,7-bis[4-(2-ethylhexyloxy)phenyl]-10H-phenothiazin-10-yl}biphenyl-4-carboxylic acid (PTZ2) were developed. Owing to their tuned and negative-shifted HOMO levels (vs. NHE), they were used as HC coadsorbents in dye-sensitized solar cells (DSSCs) to improve cell performance through desired cascade-type hole-transfer processes. Their detailed functions as HC coadsorbents in DSSCs were investigated to obtain evidence for the desired cascade-type hole-transfer processes. They have multiple functions, such as preventing π-π stacking of dye molecules, harvesting light of shorter wavelengths, and faster dye regeneration. By using PTZ2 as the tailor-made HC coadsorbent on the TiO2 surface with the organic dye NKX2677, an extremely high conversion efficiency of 8.95 % was achieved under 100â mW cm(-2) AM 1.5G simulated light (short-circuit current JSC =16.56â mA cm(-2) , open-circuit voltage VOC =740â mV, and fill factor of 73 %). Moreover, JSC was increased by 13 %, VOC by 27 % and power-conversion efficiency by 49 % in comparison to an NKX2677-based DSSC without an HC coadsorbent. This is due to the HC coadsorbent having a HOMO energy level well matched to that of the NKX-2677 dye to induce the desired cascade-type hole-transfer processes, which are associated with a slower charge recombination, fast dye regeneration, effective screening of liquid electrolytes, and an induced negative shift of the quasi-Fermi level of the electrode. Thus, this new class of Y-shaped, low molecular weight, organic, HC coadsorbents based on phenothiazine carboxylic acid derivatives hold promise for highly efficient organic DSSCs.
RESUMO
BACKGROUND: The prognosis of fulminant hepatic failure (FHF) depends on the etiology and reversibility. In this study, we identified the etiological difference of FHF in Korea and analyzed the prognostic factors after liver transplantation (LT) for FHF. METHODS: We retrospectively reviewed 42 patients with FHF who underwent LT from April 1999 to April 2011 at Samsung Medical Center, Seoul, Korea. The patients were categorized into two groups according to the short-term result of LT, and perioperative profiles were compared to identify the short-term poor prognostic factors. RESULTS: Unlike Western countries, there was no paracetamol-related FHF but herbal/folk medicines were the most frequent causes of FHF (26.2%). HAV-related FHF increased significantly and comprised the main portion of FHF with Herbal/folk medicines after 2005. Encephalopathy grade, onset time, pre-transplantation need of renal replacement, and ventilator treatment were significantly different between groups in univariate analysis. In multivariate analysis, pre-transplantation renal replacement treatment and hepatic encephalopathy grade IV were the independent prognostic factors after LT. CONCLUSIONS: The etiologies of FHF in Korea were different compared with Western reports. The requirement of renal replacement treatment and hepatic encephalopathy grade IV were identified as independent poor prognostic factors after LT for FHF in this study.
Assuntos
Falência Hepática Aguda/etiologia , Transplante de Fígado , Adolescente , Adulto , Idoso , Feminino , Humanos , Falência Hepática Aguda/diagnóstico , Falência Hepática Aguda/cirurgia , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Prognóstico , República da Coreia , Estudos Retrospectivos , Fatores de Risco , Resultado do Tratamento , Adulto JovemRESUMO
Background: Approximately 30% of diabetic patients develop diabetic nephropathy, a representative microvascular complication. Although the etiological mechanism has not yet been fully elucidated, renal tubular damage by hyperglycemia-induced expression of transforming growth factor-ß (TGF-ß) is known to be involved. Recently, a new type of cell death by iron metabolism called ferroptosis was reported to be involved in kidney damage in animal models of diabetic nephropathy, which could be induced by TGF-ß. Bone morphogenetic protein-7 (BMP7) is a well-known antagonist of TGF-ß inhibiting TGF-ß-induced fibrosis in many organs. Further, BMP7 has been reported to play a role in the regeneration of pancreatic beta cells in diabetic animal models. Methods: We used protein transduction domain (PTD)-fused BMP7 in micelles (mPTD-BMP7) for long-lasting in vivo effects and effective in vitro transduction and secretion. Results: mPTD-BMP7 successfully accelerated the regeneration of diabetic pancreas and impeded progression to diabetic nephropathy. With the administration of mPTD-BMP7, clinical parameters and representative markers of pancreatic damage were alleviated in a mouse model of streptozotocin-induced diabetes. It not only inhibited the downstream genes of TGF-ß but also attenuated ferroptosis in the kidney of the diabetic mouse and TGF-ß-stimulated rat kidney tubular cells. Conclusion: BMP7 impedes the progression of diabetic nephropathy by inhibiting the canonical TGF-ß pathway, attenuating ferroptosis, and helping regenerate diabetic pancreas.
Assuntos
Diabetes Mellitus Experimental , Nefropatias Diabéticas , Ferroptose , Animais , Camundongos , Ratos , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Pâncreas/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
DNA methylation regulates gene expression and contributes to tumorigenesis in the early stages of cancer. In colorectal cancer (CRC), CpG island methylator phenotype (CIMP) is recognized as a distinct subset that is associated with specific molecular and clinical features. In this study, we investigated the genomewide DNA methylation patterns among patients with CRC. The methylation data of 1 unmatched normal, 142 adjacent normal, and 294 tumor samples were analyzed. We identified 40,003 differentially methylated positions with 6,933 (79.8%) hypermethylated and 16,145 (51.6%) hypomethylated probes in the genic region. Hypermethylated probes were predominantly found in promoter-like regions, CpG islands, and N shore sites; hypomethylated probes were enriched in open-sea regions. CRC tumors were categorized into three CIMP subgroups, with 90 (30.6%) in the CIMP-high (CIMP-H), 115 (39.1%) in the CIMP-low (CIMP-L), and 89 (30.3%) in the non-CIMP group. The CIMP-H group was associated with microsatellite instabilityhigh tumors, hypermethylation of MLH1, older age, and rightsided tumors. Our results showed that genome-wide methylation analyses classified patients with CRC into three subgroups according to CIMP levels, with clinical and molecular features consistent with previous data. [BMB Reports 2023; 56(10): 563-568].
Assuntos
Neoplasias Colorretais , Metilação de DNA , Humanos , Metilação de DNA/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Ilhas de CpG/genética , Fenótipo , Epigênese Genética/genética , República da CoreiaRESUMO
The regulatory effect of non-coding large-scale structural variations (SVs) on proto-oncogene activation remains unclear. This study investigated SV-mediated gene dysregulation by profiling 3D cancer genome maps from 40 patients with colorectal cancer (CRC). We developed a machine learning-based method for spatial characterization of the altered 3D cancer genome. This revealed a frequent establishment of "de novo chromatin contacts" that can span multiple topologically associating domains (TADs) in addition to the canonical TAD fusion/shuffle model. Using this information, we precisely identified super-enhancer (SE)-hijacking and its clonal characteristics. Clonal SE-hijacking genes, such as TOP2B, are recurrently associated with cell-cycle/DNA-processing functions, which can potentially be used as CRC prognostic markers. Oncogene activation and increased drug resistance due to SE-hijacking were validated by reconstructing the patient's SV using CRISPR-Cas9. Collectively, the spatial and clonality-resolved analysis of the 3D cancer genome reveals regulatory principles of large-scale SVs in oncogene activation and their clinical implications.
Assuntos
Neoplasias Colorretais , Genoma , Humanos , Prognóstico , Cromatina , DNA , Neoplasias Colorretais/genéticaRESUMO
Cell-free DNA (cfDNA) sequencing has demonstrated great potential for early cancer detection. However, most large-scale studies have focused only on either targeted methylation sites or whole-genome sequencing, limiting comprehensive analysis that integrates both epigenetic and genetic signatures. In this study, we present a platform that enables simultaneous analysis of whole-genome methylation, copy number, and fragmentomic patterns of cfDNA in a single assay. Using a total of 950 plasma (361 healthy and 589 cancer) and 240 tissue samples, we demonstrate that a multifeature cancer signature ensemble (CSE) classifier integrating all features outperforms single-feature classifiers. At 95.2% specificity, the cancer detection sensitivity with methylation, copy number, and fragmentomic models was 77.2%, 61.4%, and 60.5%, respectively, but sensitivity was significantly increased to 88.9% with the CSE classifier (p value < 0.0001). For tissue of origin, the CSE classifier enhanced the accuracy beyond the methylation classifier, from 74.3% to 76.4%. Overall, this work proves the utility of a signature ensemble integrating epigenetic and genetic information for accurate cancer detection.
Assuntos
Ácidos Nucleicos Livres , Neoplasias , Humanos , Detecção Precoce de Câncer , Variações do Número de Cópias de DNA , Neoplasias/diagnóstico , Neoplasias/genética , Metilação de DNA , Biomarcadores Tumorais/genéticaRESUMO
Epiregulin (EREG) induces cell growth by binding to the epidermal growth factor receptor (EGFR). Expression of EREG affects sensitivity to cetuximab a chimeric monoclonal antibody that inhibits the EGFR signaling pathway. The mechanism through which EREG is regulated is largely unknown, but a methyl-array study previously performed by our group revealed that EREG is methylated in gastric cancer cells. In this study, we found that EREG gene expression was low in 7 out of 11 gastric cancer cells and this downregulation was mediated by aberrant CpG methylation of the EREG promoter. Treatment with 5-aza-CdR restored EREG expression and demethylated CpG sites in the EREG promoter. Compared with DNA methyltransferase 1 (DNMT1), knock-down of DNA methyltransferase 3b (DNMT3b) significantly increased the expression of EREG and led to the demethylation of specific CpG sites in the EREG promoter, suggesting that DNMT3b primarily regulates CpG methylation and silencing of the EREG gene. EREG methylation was observed in 30% (4/13) of human primary gastric tumor tissues we evaluated. In addition to DNA methylation, results from a chromatin immunoprecipitation assay demonstrated that transcriptional levels of EREG were associated with the enrichment of active histone marks (H3K4me3 and AcH3) and of a repressive mark (H3K27me2). Treatment with 5-aza-CdR dynamically increased the low occupancy of H3K4me3 and AcH3, while decreasing the high enrichment of H3K27me2, indicating that dynamic histone modifications contribute to EREG regulation in addition to DNA methylation. Finally, the combination of 5-aza-CdR and cetuximab exerted a synergistic anti-proliferative effect on gastric cancer cells. Taken together, the results of our study showed for the first time that EREG is epigenetically silenced in gastric cancer cells by aberrant DNA methylation and histone modification.
Assuntos
Fator de Crescimento Epidérmico/antagonistas & inibidores , Fator de Crescimento Epidérmico/genética , Epigênese Genética , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Humanizados , Azacitidina/administração & dosagem , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Linhagem Celular Tumoral , Cetuximab , Ilhas de CpG , DNA (Citosina-5-)-Metiltransferase 1 , DNA (Citosina-5-)-Metiltransferases/antagonistas & inibidores , DNA (Citosina-5-)-Metiltransferases/genética , Metilação de DNA , Decitabina , Epigênese Genética/efeitos dos fármacos , Epirregulina , Receptores ErbB/metabolismo , Técnicas de Silenciamento de Genes , Inativação Gênica , Histonas/genética , Histonas/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Regiões Promotoras Genéticas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , RNA Neoplásico/metabolismo , RNA Interferente Pequeno/genética , Neoplasias Gástricas/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto , DNA Metiltransferase 3BRESUMO
Ldb1 and erythroid partners SCL, GATA-1, and LMO2 form a complex that is required to establish spatial proximity between the ß-globin locus control region and gene and for transcription activation during erythroid differentiation. Here we show that Ldb1 controls gene expression at multiple levels. Ldb1 stabilizes its erythroid complex partners on ß-globin chromatin, even though it is not one of the DNA-binding components. In addition, Ldb1 is necessary for enrichment of key transcriptional components in the locus, including P-TEFb, which phosphorylates Ser2 of the RNA polymerase C-terminal domain for efficient elongation. Furthermore, reduction of Ldb1 results in the inability of the locus to migrate away from the nuclear periphery, which is necessary to achieve robust transcription of ß-globin in nuclear transcription factories. Ldb1 contributes these critical functions at both embryonic and adult stages of globin gene expression. These results implicate Ldb1 as a factor that facilitates nuclear relocation for transcription activation.