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1.
Proc Natl Acad Sci U S A ; 117(1): 584-594, 2020 01 07.
Artículo en Inglés | MEDLINE | ID: mdl-31843895

RESUMEN

In this study, we provide critical evidence that STAT2 stability regulation plays an essential role in melanoma cell proliferation and colony growth. We found that the interaction of FBXW7 and STAT2 induced STAT2 destabilization via a ubiquitination-mediated proteasomal degradation pathway. Notably, GSK3ß-mediated STAT2 phosphorylation facilitated STAT2-FBXW7 interactions via the DNA binding domain of STAT2 and domains 1, 2, 6, and 7 of FBXW7 WD40. Importantly, the inverse correlation between protein levels of STAT2 and FBXW7 were observed not only in human melanoma cells but also in a human skin cancer tissue array. The relationship between protein levels of STAT2 and FBXW7, cell proliferation, and colony growth were similarly observed in the melanoma cell lines SK-MEL-2, -5, and -28. Moreover, STAT2 knockdown in melanoma cells suppressed melanoma cell proliferation and colony formation. These data demonstrated that FBXW7-mediated STAT2 stability regulation plays an essential role in melanoma cell proliferation and cancer growth.


Asunto(s)
Proteína 7 que Contiene Repeticiones F-Box-WD/metabolismo , Melanoma/patología , Factor de Transcripción STAT2/metabolismo , Neoplasias Cutáneas/patología , Adulto , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Proliferación Celular , Femenino , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Humanos , Masculino , Persona de Mediana Edad , Fosforilación , Estabilidad Proteica , Proteolisis , Factor de Transcripción STAT2/química , Factor de Transcripción STAT2/genética , Serina/metabolismo , Transducción de Señal , Piel/patología , Treonina/metabolismo , Análisis de Matrices Tisulares , Ubiquitinación , Repeticiones WD40
2.
Int J Mol Sci ; 24(11)2023 May 31.
Artículo en Inglés | MEDLINE | ID: mdl-37298484

RESUMEN

We investigated the role of TONSL, a mediator of homologous recombination repair (HRR), in stalled replication fork double-strand breaks (DSBs) in cancer. Publicly available clinical data (tumors from the ovary, breast, stomach and lung) were analyzed through KM Plotter, cBioPortal and Qomics. Cancer stem cell (CSC)-enriched cultures and bulk/general mixed cell cultures (BCCs) with RNAi were employed to determine the effect of TONSL loss in cancer cell lines from the ovary, breast, stomach, lung, colon and brain. Limited dilution assays and ALDH assays were used to quantify the loss of CSCs. Western blotting and cell-based homologous recombination assays were used to identify DNA damage derived from TONSL loss. TONSL was expressed at higher levels in cancer tissues than in normal tissues, and higher expression was an unfavorable prognostic marker for lung, stomach, breast and ovarian cancers. Higher expression of TONSL is partly associated with the coamplification of TONSL and MYC, suggesting its oncogenic role. The suppression of TONSL using RNAi revealed that it is required in the survival of CSCs in cancer cells, while BCCs could frequently survive without TONSL. TONSL dependency occurs through accumulated DNA damage-induced senescence and apoptosis in TONSL-suppressed CSCs. The expression of several other major mediators of HRR was also associated with worse prognosis, whereas the expression of error-prone nonhomologous end joining molecules was associated with better survival in lung adenocarcinoma. Collectively, these results suggest that TONSL-mediated HRR at the replication fork is critical for CSC survival; targeting TONSL may lead to the effective eradication of CSCs.


Asunto(s)
Neoplasias , Reparación del ADN por Recombinación , Femenino , Humanos , Daño del ADN , Reparación del ADN/genética , Replicación del ADN/genética , Recombinación Homóloga , Células Madre Neoplásicas
3.
Sensors (Basel) ; 22(11)2022 Jun 06.
Artículo en Inglés | MEDLINE | ID: mdl-35684921

RESUMEN

We recently developed a long-length detector that combines three detectors and successfully acquires whole-body X-ray images. Although the developed detector system can efficiently acquire whole-body images in a short time, it may show problems with diagnostic performance in some areas owing to the use of high-energy X-rays during whole-spine and long-length examinations. In particular, during examinations of relatively thin bones, such as ankles, with a long-length detector, the image quality deteriorates because of an increase in X-ray transmission. An additional filter is primarily used to address this limitation, but this approach imposes a higher load on the X-ray tube to compensate for reductions in the radiation dose and the problem of high manufacturing costs. Thus, in this study, a newly designed additional filter was fabricated using 3D printing technology to improve the applicability of the long-length detector. Whole-spine anterior-posterior (AP), lateral, and long-leg AP X-ray examinations were performed using 3D-printed additional filters composed of 14 mm thick aluminum (Al) or 14 mm thick Al + 1 mm thick copper (Cu) composite material. The signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), and radiation dose for the acquired X-ray images were evaluated to demonstrate the usefulness of the filters. Under all X-ray inspection conditions, the most effective data were obtained when the composite additional filter based on a 14 mm thick Al + 1 mm thick Cu material was used. We confirmed that an SNR improvement of up to 46%, CNR improvement of 37%, and radiation dose reduction of 90% could be achieved in the X-ray images obtained using the composite additional filter in comparison to the images obtained with no filter. The results proved that the additional filter made with a 3D printer was effective in improving image quality and reducing the radiation dose for X-ray images obtained using a long-length detector.


Asunto(s)
Impresión Tridimensional , Fantasmas de Imagen , Dosis de Radiación , Radiografía , Relación Señal-Ruido , Rayos X
4.
Medicina (Kaunas) ; 58(7)2022 Jul 18.
Artículo en Inglés | MEDLINE | ID: mdl-35888666

RESUMEN

Background and Objectives: Prion diseases are fatal neurodegenerative disorders caused by the abnormal proteinase K-resistant prion protein (PrPSc). Since variant Creutzfeldt-Jakob disease (CJD) was first reported in the United Kingdom (UK) in 1996, the occurrence of variant CJD has been reported in over 10 countries. To date, variant CJD has not been reported in Korea. However, the E211K somatic mutation in the prion protein gene (PRNP), which is related to bovine spongiform encephalopathy (BSE), was reported in Korean Holstein cattle, and atypical BSE, which is supposed to be sporadic BSE, has been occurring in many countries, including Japan and the USA. These results suggest that BSE may occur naturally in Korea. Thus, we performed a preemptive PrPSc test in appendix specimens to diagnose variant CJD in a Korean population. Materials and Methods: In the present study, we investigated CJD-related mutations and polymorphisms of the PRNP gene and carried out an examination on PrPSc in appendix specimens of Korean patients after appendectomy. Results: In all Korean appendix specimens tested, PrPSc bands were not detected. Conclusion: To the best of our knowledge, this was the first evaluation of PrPSc in Korean appendix specimens.


Asunto(s)
Apéndice , Síndrome de Creutzfeldt-Jakob , Encefalopatía Espongiforme Bovina , Enfermedades por Prión , Priones , Animales , Apéndice/metabolismo , Bovinos , Síndrome de Creutzfeldt-Jakob/genética , Síndrome de Creutzfeldt-Jakob/metabolismo , Encefalopatía Espongiforme Bovina/metabolismo , Endopeptidasa K , Enfermedades por Prión/genética , Proteínas Priónicas/genética , Priones/genética , Priones/metabolismo
5.
Ann Hum Genet ; 85(2): 80-91, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33249558

RESUMEN

Phosphodiesterase 3A (PDE3A) is an enzyme that plays an important role in the regulation of cyclic adenosine monophosphate (cAMP)-mediated intracellular signaling in cardiac myocytes and platelets. PDE3A hydrolyzes cAMP, which results in a decrease in intracellular cAMP levels and leads to platelet activation. Whole-exome sequencing of 50 DNA samples from a healthy Korean population revealed a total of 13 single nucleotide polymorphisms including five missense variants, D12N, Y497C, H504Q, C707R, and A980V. Recombinant proteins for the five variants of PDE3A (and wild-type protein) were expressed in a FreeStyle 293 expression system with site-directed mutagenesis. The expression of the recombinant PDE3A proteins was confirmed with Western blotting. Catalytic activity of the PDE3A missense variants and wild-type enzyme was measured with a PDE-based assay. Effects of the missense variants on the inhibition of PDE3A activity by cilostazol were also investigated. All variant proteins showed reduced activity (33-53%; p < .0001) compared to the wild-type protein. In addition, PDE3A activity was inhibited by cilostazol in a dose-dependent manner and was further suppressed in the missense variants. Specifically, the PDE3A Y497C showed significantly reduced activity, consistent with the predictions of in silico analyses. The present study provides evidence that individuals carrying the PDE3A Y497C variant may have lower enzyme activity for cAMP hydrolysis, which could cause interindividual variation in cAMP-mediated physiological functions.


Asunto(s)
Cilostazol/administración & dosificación , AMP Cíclico/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 3/genética , Activación Plaquetaria/efectos de los fármacos , Adulto , Plaquetas/efectos de los fármacos , Cilostazol/efectos adversos , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Masculino , Mutación Missense/genética , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Activación Plaquetaria/genética , Inhibidores de Agregación Plaquetaria/administración & dosificación , Inhibidores de Agregación Plaquetaria/efectos adversos , Polimorfismo Genético/genética , Transducción de Señal/efectos de los fármacos , Secuenciación del Exoma
6.
J Nat Prod ; 84(2): 298-309, 2021 02 26.
Artículo en Inglés | MEDLINE | ID: mdl-33529025

RESUMEN

Biological species collections are critical for natural product drug discovery programs. However, prioritization of target species in massive collections remains difficult. Here, we introduce an untargeted metabolomics-based prioritization workflow that uses MS/MS molecular networking to estimate scaffold-level distribution. As a demonstration, we applied the workflow to 40 polyporoid fungal species. Nine species were prioritized as candidates based on the chemical structural and compositional similarity (CSCS) metric. Most of the selected species showed relatively higher richness and uniqueness of metabolites than those of the others. Cryptoporus volvatus, one of the prioritized species, was investigated further. The chemical profiles of the extracts of C. volvatus culture and fruiting bodies were compared, and it was shown that derivative-level diversity was higher in the fruiting bodies; meanwhile, scaffold-level diversity was similar. This showed that the compounds found from a cultured fungus can also be isolated in wild mushrooms. Targeted isolation of the fruiting body extract yielded three unknown (1-3) and six known (4-9) cryptoporic acid derivatives, which are drimane-type sesquiterpenes with isocitric acid moieties that have been reported in this species. Cryptoporic acid T (1) is a trimeric cryptoporic acid reported for the first time. Compounds 2 and 5 exhibited cytotoxicity against HCT-116 cell lines with IC50 values of 4.3 and 3.6 µM, respectively.


Asunto(s)
Isocitratos/aislamiento & purificación , Sesquiterpenos Policíclicos/aislamiento & purificación , Polyporaceae/química , Polyporaceae/clasificación , Cuerpos Fructíferos de los Hongos/química , Células HCT116 , Humanos , Estructura Molecular , Sesquiterpenos Policíclicos/farmacología , República de Corea , Espectrometría de Masas en Tándem
7.
Odontology ; 108(2): 194-201, 2020 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-31512103

RESUMEN

The purpose of this study is to assess the effect of autoclave sterilization on the cyclic fatigue and torsional fracture resistance of ProTaper Universal (PTU), K3XF, HyFlex EDM (EDM), and TF adaptive (TFA). Sixty instruments from each file type were divided into two categories for cyclic fatigue group (CGr) and torsional fracture group (TGr). CGr and TGr were divided into three subgroups, respectively, consisting of ten instruments from each file type. Cyclic fatigue fracture test was performed using artificial canal made of stainless steel, and the mean number of cycles to failure (NCF) were determined. CGr1, the files were tested to establish baseline for NCF; CGr2, the files were tested cyclic fatigue after 10 cycles of autoclave; CGr3, instruments were autoclaved after being cycled to 25, 50, and 75% of corresponding NCF determined in CGr1, followed by cyclic fatigue test. The torsional fracture test was performed without autoclave (TGr1), after 3-cycle autoclave (TGr2), and 7-cycle autoclave (TGr3), respectively, which evaluated maximum torque and angular deflection. NCF, maximum torque and angular deflection were compared using one-way ANOVA with Bonferroni test. Two-way ANOVA was performed to determine the interaction between 'autoclave treatment' and 'type of NiTi file'. EDM showed highest NCF within the same autoclave treatment. TFA presented the lowest maximum torque and the highest angular deflection, and PTU presented the lowest angular deflection. Within the same NiTi file systems, most of NCF, maximum torque and angular deflection of tested files were not significantly influenced by autoclave condition.


Asunto(s)
Instrumentos Dentales , Preparación del Conducto Radicular , Aleaciones Dentales , Diseño de Equipo , Falla de Equipo , Ensayo de Materiales , Esterilización , Estrés Mecánico , Titanio
8.
Biochem Biophys Res Commun ; 519(1): 100-105, 2019 10 29.
Artículo en Inglés | MEDLINE | ID: mdl-31481234

RESUMEN

Lipid metabolism is associated with colon cancer prognosis and incidence. Stearoyl-CoA desaturase 1 (SCD1), which converts fully saturated fatty acids (SFAs) to monounsaturated fatty acids (MUFAs), has been suggested as a vulnerable target for selective elimination of cancer stem cells (CSCs). However, the clinical significance and physiological role of SCD1 in CSCs has not been well demonstrated. Here, we showed the clinical and biochemical relevance of blocking SCD1 to target CSCs by analyzing human colon cancer data from TCGA and through lipidomic profiling of CSCs with or without SCD1 inhibition using mass spectrometry. Positive associations between SCD1 expression and colorectal cancer patient clinical status and the expression of CSC-related genes (WNT and NOTCH signaling) were found based on TCGA data analysis. Lipidomic profiling of CSCs and bulk cancer cells (BCCs) using mass spectrometry revealed that colon CSCs contained a distinctive lipid profile, with higher free MUFA and lower free SFA levels than in BCCs, suggesting that enhanced SCD1 activity generates MUFAs that may support WNT signaling in CSCs. In addition, all identified phosphatidyl-ethanolamine-containing MUFAs were found at higher levels in CSCs. Interestingly, we observed lower phosphatidyl-serine (18:1/18:0), phosphatidyl-choline (PC; p-18:0/18:1)), and sphingomyelin (SM; d18:1/20:0 or d16:1/22:0) levels in CSCs than in BCCs. Of those, SCD1 inhibition, which efficiently diminished free MUFA levels, increased those specific PC and SM and MUFAs in CSCs promptly. These results suggest that these specific lipid composition is critical for CSC stem cell maintenance. In addition, not only free MUFAs, which are known to be required for WNT signaling, but also other phospholipids, such as SM, which are important for lipid raft formation, may mediate other cell signaling pathways that support CSC maintenance. Comparison of the lipidomic profiles of colon cancer cells with those of previously reported for glioma cells further demonstrated the tissue specific characteristics of lipid metabolism in CSCs.


Asunto(s)
Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Ácidos Grasos Monoinsaturados/metabolismo , Células Madre Neoplásicas/metabolismo , Línea Celular Tumoral , Neoplasias del Colon/genética , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica , Humanos , Metabolismo de los Lípidos , Células Madre Neoplásicas/patología , Fosfolípidos/metabolismo , Transducción de Señal , Estearoil-CoA Desaturasa/genética , Estearoil-CoA Desaturasa/metabolismo
9.
Molecules ; 24(24)2019 Dec 10.
Artículo en Inglés | MEDLINE | ID: mdl-31835579

RESUMEN

Evodiamine, an alkaloid contained in traditional Asian herbal medicines that have been used for hundreds years, is interesting due to its cytotoxic effects against many cancers. We examined the effect of evodiamine on the cancer stem cell (CSC) population and the bulk cultured cancer cells (BCC) of colon cancers to examine the double targeting effect. We found that three colon cancer cell lines' BCC and CSC are effectively targeted by evodiamine. Evodiamine was able to suppress BCC proliferation and induce apoptosis of the cells captured in G2/M phase, as previously reported. However, evodiamine did not cause the accumulation of CSCs at a certain stage of the cell cycle, resulting in the elimination of stemness through an unknown mechanism. By analyzing the expression of 84 genes related to CSCs in two colon cancer cell lines' CSC, as well as performing further informatics analyses, and quantitative RT-PCR analyses of 24 CSC genes, we found that evodiamine suppressed the expression of the genes that control key signaling pathways of CSC, namely, WNT and NOTCH signaling, to lead CSC elimination. These results suggest that evodiamine should be further developed for targeting both BCCs and CSCs in colon cancers.


Asunto(s)
Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Quinazolinas/farmacología , Receptores Notch/metabolismo , Transducción de Señal/efectos de los fármacos , Proteínas Wnt/metabolismo , Antineoplásicos/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Células Madre Neoplásicas/patología
10.
FASEB J ; 31(2): 625-635, 2017 02.
Artículo en Inglés | MEDLINE | ID: mdl-27811063

RESUMEN

Cancer stem-like cells (CSLCs) contribute to the initiation and recurrence of tumors and to their resistance to conventional therapies. In this study, small interfering RNA (siRNA)-based screening of ∼4800 druggable genes in 3-dimensional CSLC cultures in comparison to 2-dimensional bulk cultures of U87 glioma cells revealed 3 groups of genes essential for the following: survival of the CSLC population only, bulk-cultured population only, or both populations. While diverse biologic processes were associated with siRNAs reducing the bulk-cultured population, CSLC-eliminating siRNAs were enriched in a few functional categories, such as lipid metabolism, protein metabolism, and gene expression. Interestingly, siRNAs that selectively reduced CSLC only were found to target genes for cholesterol and unsaturated fatty acid synthesis. The lipidomic profile of CSLCs revealed increased levels of monounsaturated lipids. Pharmacologic blockage of these target pathways reduced CSLCs, and this effect was eliminated by addition of downstream metabolite products. The present CSLC-sensitive target categories provide a useful resource that can be exploited for the selective elimination of CSLCs.-Song, M., Lee, H., Nam, M.-H., Jeong, E., Kim, S., Hong, Y., Kim, N., Yim, H. Y., Yoo, Y.-J., Kim, J. S., Kim, J.-S., Cho, Y.-Y., Mills, G. B., Kim, W.-Y., Yoon, S. Loss-of-function screens of druggable targetome against cancer stem-like cells.


Asunto(s)
Regulación Neoplásica de la Expresión Génica/fisiología , Células Madre Neoplásicas/efectos de los fármacos , Animales , Línea Celular , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones SCID , Neoplasias Experimentales/metabolismo , Interferencia de ARN , ARN Interferente Pequeño
11.
BMC Bioinformatics ; 18(Suppl 12): 423, 2017 Oct 16.
Artículo en Inglés | MEDLINE | ID: mdl-29072139

RESUMEN

BACKGROUND: A network motif is defined as a statistically significant and recurring subgraph pattern within a network. Most existing instance collection methods are not feasible due to high memory usage issues and provision of limited network motif information. They require a two-step process that requires network motif identification prior to instance collection. Due to the impracticality in obtaining motif instances, the significance of their contribution to problem solving is debated within the field of biology. RESULTS: This paper presents NemoProfile, an efficient new network motif data model. NemoProfile simplifies instance collection by resolving memory overhead issues and is seamlessly generated, thus eliminating the need for costly two-step processing. Additionally, a case study was conducted to demonstrate the application of network motifs to existing problems in the field of biology. CONCLUSION: NemoProfile comprises network motifs and their instances, thereby facilitating network motifs usage in real biological problems.


Asunto(s)
Algoritmos , Biología Computacional/métodos , Redes Reguladoras de Genes , Mapas de Interacción de Proteínas , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo
12.
Chem Pharm Bull (Tokyo) ; 65(4): 349-355, 2017 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-28132960

RESUMEN

Many aminodihydroquinoline compounds have been studied to determine their cytotoxicity to cancer cells. However, anti-cancer stem cells (CSCs) activity of aminodihydroquinoline has not been tested in spite that CSC is believed to do an important roles in chemotherapy resistance and recurrence. The CSC selective targeting activities of 10 recently synthesized 2-aminodihydroquinoline analogs were examined on CSCs and bulk culture of a glioblastoma cell line. A diethylaminopropyl substituted aminodihydroquinoline, 5h, showed a strong anti-CSC effect and general cytotoxicity. However, a benzyl substituted aminodihydroquinoline, 5i, displayed the most effective anti-CSC effect, with no or small significant cytotoxic effect in bulk culture conditions. While 5h temporarily enhanced CSC marker-positive cells and eventually suppressed the CSC population, which is similar to other cytotoxic anticancer reagents reported, 5i selectively eliminated CSC marker-positive cells based on fluorescence activated cell sorter (FACS) analysis. 5h also temporarily activated some genes associated with signaling required for CSC, while 5i selectively suppressed these genes supporting that the differential effects are resulted from different molecular responses. In addition, the selective CSC effect is also found against a colon cancer cell line. Collectively, we suggest that these two novel aminodihydroquinoline compounds possess novel anti-CSC effects in colon and brain tumor derived cell lines probably through independent pathways.


Asunto(s)
Antineoplásicos/farmacología , Células Madre Neoplásicas/efectos de los fármacos , Quinolinas/farmacología , Antineoplásicos/química , Relación Dosis-Respuesta a Droga , Humanos , Estructura Molecular , Quinolinas/química , Relación Estructura-Actividad , Células Tumorales Cultivadas
13.
Biochem Biophys Res Commun ; 469(4): 1153-8, 2016 Jan 22.
Artículo en Inglés | MEDLINE | ID: mdl-26713361

RESUMEN

In spite of the recent improvements, the resistance to chemotherapy/radiotherapy followed by relapse is the main hurdle for the successful treatment of breast cancer, a leading cause of death in women. A small population of breast cancer cells that have stem-like characteristics (cancer stem-like cells; CSLC) may contribute to this resistance and relapse. Here, we report on a component of a traditional Chinese medicine, evodiamine, which selectively targets CSLC of breast cancer cell lines MCF7 and MDAMB 231 at a concentration that does show a little or no cytotoxic effect on bulk cancer cells. While evodiamine caused the accumulation of bulk cancer cells at the G2/M phase, it did not hold CSLC in a specific cell cycle phase but instead, selectively killed CSLC. This was not due to the culture of CSLC in suspension or without FBS. A proteomic analysis and western blotting revealed that evodiamine changed the expression of cell cycle regulating molecules more efficiently in CSLC cells than in bulk cancer cells. Surprisingly, evodiamine selectively activated p53 and p21 and decreased inactive Rb, the master molecules in G1/S checkpoint. These data collectively suggest a novel mechanism involving CSLC-specific targeting by evodiamine and its possible use to the therapy of breast cancer.


Asunto(s)
Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Células Madre Neoplásicas/metabolismo , Quinazolinas/administración & dosificación , Proteína de Retinoblastoma/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Antineoplásicos Fitogénicos/administración & dosificación , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Células MCF-7 , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/patología , Transducción de Señal/efectos de los fármacos , Resultado del Tratamiento
14.
Biochem Biophys Res Commun ; 470(2): 294-299, 2016 Feb 05.
Artículo en Inglés | MEDLINE | ID: mdl-26774343

RESUMEN

Growing evidence shows that cancer stem-like cells (CSLCs) contribute to breast cancer recurrence and to its resistance to conventional therapies. The extracellular signal-regulated kinase (ERK) signaling pathway is a major determinant in the control of diverse cellular processes, including the maintenance of CSLCs. In this study, we found that Kazinol-E, an antioxidant flavan from Broussonetia kazinoki, decreased the CSLC population of a breast cancer cell line, MCF7. The CSLC population, characterized by CD44 high/CD24 low expression or by high Aldehyde dehydrogenase 1 activity, was decreased by a concentration of Kazinol-E that did not affect the growth of bulk-cultured MCF7 cells. Kazinol-E did not decrease EGF-induced ERK phosphorylation in CSLCs, but did block the phosphorylation of an ERK substrate, p90RSK2, at Thr359/Ser363. We further demonstrated that EGF-induced ERK activity was blocked by Kazinol-E in a wild-type K-Ras-expressing non-small cell lung cancer cell line H226B. An in vitro kinase assay with purified ERK1 and p90RSK2 as its substrate demonstrated a direct inhibition of ERK activity by Kazinol E. Additionally, a the molecular docking study provided putative binding modes of Kazinol-E into the ATP binding pocket of ERK1 Collectively, these results suggest that Kazinol-E is a direct inhibitor of ERK1, and more studies are warranted to develop this reagent for therapeutic breast CSLC targeting.


Asunto(s)
Neoplasias de la Mama/enzimología , Neoplasias de la Mama/patología , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Flavonoides/administración & dosificación , Células Madre Neoplásicas/enzimología , Células Madre Neoplásicas/patología , Neoplasias de la Mama/tratamiento farmacológico , Carcinogénesis/efectos de los fármacos , Carcinogénesis/patología , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células MCF-7 , Células Madre Neoplásicas/efectos de los fármacos
15.
Bioorg Med Chem Lett ; 26(13): 3067-3072, 2016 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-27209235

RESUMEN

A novel series of benzoxazole analogs was designed and synthesized, and their inhibitory activities against Aurora kinases were evaluated. Some of the tested compounds exhibited a promising activity with respect to the inhibition of Aurora B kinase. A structure-activity relationship study indicated that linker length, regiochemistry, and halogen substitution play important roles in kinase inhibitory potency. The binding modes between representative compounds and Aurora kinases were interpreted through a molecular docking study to explain the inhibitory activity and selectivity for Aurora A and B kinases. Compounds 13l and 13q also show an antiproliferative effect on the human tumor cell lines in a dose-dependent manner. The most potent 13q demonstrated good efficacy in the prostate cancer PC-3 tumor xenograft model.


Asunto(s)
Antineoplásicos/farmacología , Aurora Quinasa B/antagonistas & inhibidores , Benzoxazoles/farmacología , Diseño de Fármacos , Neoplasias Experimentales/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Aurora Quinasa B/metabolismo , Benzoxazoles/síntesis química , Benzoxazoles/química , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Ratones , Ratones Desnudos , Modelos Moleculares , Estructura Molecular , Neoplasias Experimentales/patología , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Relación Estructura-Actividad
16.
J Biol Chem ; 289(7): 3901-12, 2014 Feb 14.
Artículo en Inglés | MEDLINE | ID: mdl-24302738

RESUMEN

The target of rapamycin (TOR) kinase pathway regulates various biological processes, including translation, synthesis of ribosomal proteins, and transcription of rRNA. The ribosomal protein S6 (RPS6) is one of the well known downstream components of the TOR pathway. Ribosomal proteins have been known to have diverse functions in regulating cellular metabolism as well as protein synthesis. So far, however, little is known about other possible role(s) of RPS6 in plants, besides being a component of the 40 S ribosomal subunit and acting as a target of TOR. Here, we report that RPS6 may have a novel function via interaction with histone deacetylase 2B (AtHD2B) that belongs to the plant-specific histone deacetylase HD2 family. RPS6 and AtHD2B were localized to the nucleolus. Co-expression of RPS6 and AtHD2B caused a change in the location of both RPS6 and AtHD2B to one or several nucleolar spots. ChIP analysis suggests that RPS6 directly interacts with the rRNA gene promoter. Protoplasts overexpressing both AtHD2B and RPS6 exhibited down-regulation of pre-18 S rRNA synthesis with a concomitant decrease in transcription of some of the ribosomal proteins, suggesting their direct role in ribosome biogenesis and plant development. This is consistent with the mutation in rps6b that results in reduction in 18 S rRNA transcription and decreased root growth. We propose that the interaction between RPS6 and AtHD2B brings about a change in the chromatin structure of rDNA and thus plays an important role in linking TOR signaling to rDNA transcription and ribosome biogenesis in plants.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Nucléolo Celular/metabolismo , Genes de Plantas/fisiología , Genes de ARNr/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , ARN de Planta/metabolismo , ARN Ribosómico 18S/biosíntesis , Arabidopsis/citología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Nucléolo Celular/genética , ADN de Plantas/genética , ADN de Plantas/metabolismo , ADN Ribosómico/genética , ADN Ribosómico/metabolismo , Epigénesis Genética/fisiología , Histona Desacetilasas/genética , Histona Desacetilasas/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Regiones Promotoras Genéticas/fisiología , Protoplastos/citología , Protoplastos/metabolismo , ARN de Planta/genética , ARN Ribosómico 18S/genética , Transcripción Genética/fisiología
17.
Biochem Biophys Res Commun ; 465(2): 200-5, 2015 Sep 18.
Artículo en Inglés | MEDLINE | ID: mdl-26241676

RESUMEN

The ribosomal protein S6 (RPS6) is a downstream component of the signaling mediated by the target of rapamycin (TOR) kinase that acts as a central regulator of the key metabolic processes, such as protein translation and ribosome biogenesis, in response to various environmental cues. In our previous study, we identified a novel role of plant RPS6, which negatively regulates rDNA transcription, forming a complex with a plant-specific histone deacetylase, AtHD2B. Here we report that the Arabidopsis RPS6 interacts additionally with a histone chaperone, nucleosome assembly protein 1(AtNAP1;1). The interaction does not appear to preclude the association of RPS6 with AtHD2B, as the AtNAP1 was also able to interact with AtHD2B as well as with an RPS6-AtHD2B fusion protein in the BiFC assay and pulldown experiment. Similar to a positive effect of the ribosomal S6 kinase 1 (AtS6K1) on rDNA transcription observed in this study, overexpression or down regulation of the AtNAP1;1 resulted in concomitant increase and decrease, respectively, in rDNA transcription suggesting a positive regulatory role played by AtNAP1 in plant rDNA transcription, possibly through derepression of the negative effect of the RPS6-AtHD2B complex.


Asunto(s)
Adenosina Trifosfatasas/genética , Proteínas de Arabidopsis/genética , Arabidopsis/genética , ADN Ribosómico/genética , Histona Desacetilasas/genética , Transcripción Genética , Adenosina Trifosfatasas/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Sitios de Unión , ADN Ribosómico/metabolismo , Regulación de la Expresión Génica de las Plantas , Genes Reporteros , Histona Desacetilasas/metabolismo , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Nucleosomas/genética , Nucleosomas/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Unión Proteica , Protoplastos/metabolismo , Ribosomas/genética , Ribosomas/metabolismo , Transducción de Señal
18.
New Phytol ; 205(1): 316-28, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25187356

RESUMEN

Arabidopsis thaliana homeobox 12 (ATHB12), a homeodomain-leucine zipper class I (HD-Zip I) gene, is highly expressed in leaves and stems, and induced by abiotic stresses, but its role in development remains obscure. To understand its function during plant development, we studied the effects of loss and gain of function. Expression of ATHB12 fused to the EAR-motif repression domain (SRDX) - P35 S ::ATHB12SRDX (A12SRDX) and PATHB 12 ::ATHB12SRDX - slowed both leaf and root growth, while the growth of ATHB12-overexpressing seedlings (A12OX) was accelerated. Microscopic examination revealed changes in the size and number of leaf cells. Ploidy was reduced in A12SRDX plants, accompanied by decreased cell expansion and increased cell numbers. By contrast, cell size was increased in A12OX plants, along with increased ploidy and elevated expression of cell cycle switch 52s (CCS52s), which are positive regulators of endoreduplication, indicating that ATHB12 promotes leaf cell expansion and endoreduplication. Overexpression of ATHB12 led to decreased phosphorylation of Arabidopsis thaliana ribosomal protein S6 (AtRPS6), a regulator of cell growth. In addition, induction of ATHB12 in the presence of cycloheximide increased the expression of several genes related to cell expansion, such as EXPANSIN A10 (EXPA10) and DWARF4 (DWF4). Our findings strongly suggest that ATHB12 acts as a positive regulator of endoreduplication and cell growth during leaf development.


Asunto(s)
Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Endorreduplicación , Leucina Zippers , Hojas de la Planta/citología , Hojas de la Planta/crecimiento & desarrollo , Arabidopsis/citología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proliferación Celular/efectos de los fármacos , Forma de la Célula/efectos de los fármacos , Pared Celular/efectos de los fármacos , Pared Celular/genética , ADN de Plantas/genética , Endorreduplicación/efectos de los fármacos , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Genes de Plantas , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Fenotipo , Fosforilación/efectos de los fármacos , Hojas de la Planta/efectos de los fármacos , Raíces de Plantas/citología , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Plantas Modificadas Genéticamente , Ploidias
19.
Arch Biochem Biophys ; 569: 10-8, 2015 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-25623425

RESUMEN

Prostacyclin synthase (CYP8A1) is an enzyme responsible for the biosynthesis of prostacyclin (PGI2) which inhibits platelet activation and exhibits anti-inflammatory effect. The objectives of this study were to identify CYP8A1 genetic variants and characterize functional consequences of CYP8A1 variants. In total, 27 variants including four previously unidentified single-nucleotide polymorphisms (SNPs) were identified by direct DNA sequencing in Koreans (n=48). Among them, CYP8A1 A447T and E314Stop were newly assigned as CYP8A1(∗)5 and CYP8A1(∗)6 by the Human Cytochrome P450 Allele Nomenclature Committee, respectively. CYP8A1(∗)5 was found in the heme binding area in three individuals as a heterozygous mutation. To investigate the functional change of CYP8A1(∗)5, CYP8A1(∗)5 and wild-type CYP8A1 protein were overexpressed in an Escherichia coli expression system and purified. Metabolism of PGH2 by the CYP8A1(∗)5 protein exhibited significantly decreased activity, resulting in a 45% decrease in Vmax and a 1.8-fold decrease in intrinsic clearance compared to the wild-type. Based on the predicted crystal structure of CYP8A1(∗)5 using the Molecular Operating Environment platform, the distance from CYP8A1 Cys441 to the heme was altered with a significantly changed binding free energy for the mutant protein. Further studies would be needed to determine the effect of CYP8A1(∗)5 on PGI2 levels in humans.


Asunto(s)
Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Variación Genética , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Secuencia de Bases , Sitios de Unión/genética , Codón sin Sentido , Simulación por Computador , Sistema Enzimático del Citocromo P-450/química , ADN/genética , Hemo/metabolismo , Heterocigoto , Humanos , Cinética , Desequilibrio de Ligamiento , Modelos Moleculares , Datos de Secuencia Molecular , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Mutación , Polimorfismo de Nucleótido Simple , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , República de Corea , Homología de Secuencia de Aminoácido
20.
Carcinogenesis ; 35(10): 2232-43, 2014 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-24942865

RESUMEN

Insulin-like growth factor (IGF)-dependent and -independent antitumor activities of insulin-like growth factor binding protein-3 (IGFBP-3) have been proposed in human non-small cell lung cancer (NSCLC) cells. However, the mechanism underlying regulation of IGFBP-3 expression in NSCLC cells is not well understood. In this study, we show that activation of Akt, especially Akt3, plays a major role in the mRNA expression and protein stability of IGFBP-3 and thus antitumor activities of IGFBP-3 in NSCLC cells. When Akt was activated by genomic or pharmacologic approaches, IGFBP-3 transcription and protein stability were decreased. Conversely, suppression of Akt increased IGFBP-3 mRNA levels and protein stability in NSCLC cell lines. Characterization of the effects of constitutively active form of each Akt subtype (HA-Akt-DD) on IGFBP-3 expression in NSCLC cells and a xenograft model indicated that Akt3 plays a major role in the Akt-mediated regulation of IGFBP-3 expression and thus suppression of Akt effectively enhances the antitumor activities of IGFBP-3 in NSCLC cells with Akt3 overactivation. Collectively, these data suggest a novel function of Akt3 as a negative regulator of IGFBP-3, indicating the possible benefit of a combined inhibition of IGFBP-3 and Akt3 for the treatment of patients with NSCLC.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/genética , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Neoplasias Pulmonares/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Núcleo Celular/genética , Núcleo Celular/metabolismo , Cromonas/farmacología , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Ratones , Ratones Desnudos , Morfolinas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Estabilidad Proteica , Proteínas Proto-Oncogénicas c-akt/genética , Transcripción Genética , Ensayos Antitumor por Modelo de Xenoinjerto
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