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CD73 is a cell-surface ectoenzyme that hydrolyzes the conversion of extracellular adenosine monophosphate to adenosine, which in turn can promote resistance to immune checkpoint blockade therapy. Immune response may therefore be improved by targeting tumor CD73, and this possibility underlines the need to non-invasively assess tumor CD73 level. In this study, we developed a cysteine site-specific 89Zr-labeled anti-CD73 (89Zr-CD73) IgG immuno-PET technique that can image tumor CD73 expression in living bodies. Anti-CD73 IgG was reduced with tris(2-carboxyethyl)phosphine, underwent sulfohydryl moiety-specific conjugation with deferoxamine-maleimide, and was radiolabeled with 89Zr. CT26 mouse colon cancer cells, CT26/CD73 cells engineered to constitutively overexpress CD73, and 4T1.2 mouse breast cancer cells underwent cell binding assays and western blotting. Balb/c nude mice bearing tumors underwent 89Zr-CD73 IgG PET imaging and biodistribution studies. 89Zr-CD73 IgG showed 20-fold higher binding to overexpressing CT26/CD73 cells compared to low-expressing CT26 cells, and moderate expressing 4T1.2 cells showed uptake that was 38.9 ± 1.51% of CT26/CD73 cells. Uptake was dramatically suppressed by excess unlabeled antibody. CD73 content proportionately increased in CT26 and CT26/CD73 cell mixtures was associated with linear increases in 89Zr-CD73 IgG uptake. 89Zr-CD73 IgG PET/CT displayed clear accumulation in CT26/CD73 tumors with greater uptake compared to CT26 tumors (3.13 ± 1.70%ID/g vs. 1.27 ± 0.31%ID/g at 8 days; P = 0.04). Specificity was further supported by low CT26/CD73 tumor-to-blood ratio of 89Zr-isotype-IgG compared to 89Zr-CD73 IgG (0.48 ± 0.08 vs. 2.68 ± 0.52 at 4 days and 0.53 ± 0.07 vs. 4.81 ± 1.02 at 8 days; both P < 0.001). Immunoblotting and immunohistochemistry confirmed strong CD73 expression in CT26/CD73 tumors and low expression in CT26 tumors. 4T1.2 tumor mice also showed clear 89Zr-CD73 IgG accumulation at 8 days (3.75 ± 0.70%ID/g) with high tumor-to-blood ratio compared to 89Zr-isotype-IgG (4.91 ± 1.74 vs. 1.20 ± 0.28; P < 0.005). 89Zr-CD73 IgG specifically targeted CD73 on high expressing cancer cells in vitro and tumors in vivo. Thus, 89Zr-CD73 IgG immuno-PET may be useful for the non-invasive monitoring of CD73 expression in tumors of living subjects.
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5'-Nucleotidase , Neoplasias do Colo , Cisteína , Tomografia por Emissão de Pósitrons , Zircônio , Animais , 5'-Nucleotidase/metabolismo , Zircônio/química , Neoplasias do Colo/diagnóstico por imagem , Neoplasias do Colo/metabolismo , Neoplasias do Colo/imunologia , Neoplasias do Colo/patologia , Camundongos , Linhagem Celular Tumoral , Tomografia por Emissão de Pósitrons/métodos , Cisteína/metabolismo , Humanos , Radioisótopos , Feminino , Camundongos Endogâmicos BALB C , Distribuição Tecidual , Camundongos Nus , Proteínas Ligadas por GPI/metabolismo , Proteínas Ligadas por GPI/imunologia , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismoRESUMO
Very late antigen-4 (VLA4; CD49d) is a promising immune therapy target in treatment-resistant leukemia and multiple myeloma, and there is growing interest in repurposing the humanized monoclonal antibody (Ab), natalizumab, for this purpose. Positron emission tomography with radiolabeled Abs (immuno-PET) could facilitate this effort by providing information on natalizumab's in vivo pharmacokinetic and target delivery properties. In this study, we labeled natalizumab with 89Zr specifically on sulfhydryl moieties via maleimide-deferoxamine conjugation. High VLA4-expressing MOLT4 human T cell acute lymphoblastic leukemia cells showed specific 89Zr-natalizumab binding that was markedly blocked by excess Ab. In nude mice bearing MOLT4 tumors, 89Zr-natalizumab PET showed high-contrast tumor uptake at 7 days postinjection. Biodistribution studies confirmed that uptake was the highest in MOLT4 tumors (2.22 ± 0.41%ID/g) and the liver (2.33 ± 0.76%ID/g), followed by the spleen (1.51 ± 0.42%ID/g), while blood activity was lower at 1.12 ± 0.21%ID/g. VLA4-specific targeting in vivo was confirmed by a 58.1% suppression of tumor uptake (0.93 ± 0.15%ID/g) when excess Ab was injected 1 h earlier. In cultured MOLT4 cells, short-term 3 day exposure to the proteasome inhibitor bortezomib (BTZ) did not affect the α4 integrin level, but BTZ-resistant cells that survived the treatment showed increased α4 integrin expression. When the effects of BTZ treatment were tested in mice, there was no change of the α4 integrin level or 89Zr-natalizumab uptake in MOLT4 leukemia tumors, which underscores the complexity of tumor VLA4 regulation in vivo. In conclusion, 89Zr-natalizumab PET may be useful for noninvasive monitoring of tumor VLA4 and may assist in a more rational application of Ab-based therapies for hematologic malignancies.
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Integrina alfa4beta1 , Leucemia , Humanos , Animais , Camundongos , Natalizumab/uso terapêutico , Cisteína , Integrina alfa4 , Camundongos Nus , Distribuição Tecidual , Linhagem Celular Tumoral , Tomografia por Emissão de Pósitrons/métodos , Zircônio/químicaRESUMO
Although deep learning architecture has been used to process sequential data, only a few studies have explored the usefulness of deep learning algorithms to detect glaucoma progression. Here, we proposed a bidirectional gated recurrent unit (Bi-GRU) algorithm to predict visual field loss. In total, 5413 eyes from 3321 patients were included in the training set, whereas 1272 eyes from 1272 patients were included in the test set. Data from five consecutive visual field examinations were used as input; the sixth visual field examinations were compared with predictions by the Bi-GRU. The performance of Bi-GRU was compared with the performances of conventional linear regression (LR) and long short-term memory (LSTM) algorithms. Overall prediction error was significantly lower for Bi-GRU than for LR and LSTM algorithms. In pointwise prediction, Bi-GRU showed the lowest prediction error among the three models in most test locations. Furthermore, Bi-GRU was the least affected model in terms of worsening reliability indices and glaucoma severity. Accurate prediction of visual field loss using the Bi-GRU algorithm may facilitate decision-making regarding the treatment of patients with glaucoma.
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Glaucoma , Campos Visuais , Humanos , Reprodutibilidade dos Testes , Olho , Algoritmos , Glaucoma/diagnósticoRESUMO
RIP1 (receptor-interacting protein kinase 1) is an important component of TNF-α signaling that contributes to various pathological effects. Here, we revealed new potential roles of RIP1 in controlling WNT/ß-catenin canonical signaling to enhance metastasis of colorectal cancer (CRC). First, we showed that WNT3A treatment sequentially increased the expression of RIP1 and ß-catenin. Immunohistochemical analyses of human CRC tissue arrays consisting of normal, primary, and metastatic cancers indicated that elevated RIP1 expression might be related to ß-catenin expression, carcinogenesis, and metastasis. Intravenous injection of RIP1 over-expressed CRC cells into mice has demonstrated that RIP1 may promote metastasis. Immunoprecipitation (IP) results indicated that WNT3A treatment induces direct binding between RIP1 and ß-catenin, and that this stabilizes the ß-catenin protein in a manner that depends on the regulation of RIP1 ubiquitination via downregulation of the E3 ligase, cIAP1/2. Elimination of cIAP1/2 expression and inhibition of its ubiquitinase activity enhance WNT3A-induced RIP1 and ß-catenin protein expression and binding, which stimulates endothelial-mesenchymal transition (EMT) induction to enhance the migration and invasion of CRC cells in vitro. The results of the in vitro binding assay and IP of exogenous RIP1-containing CRC cells additionally verified the direct binding of RIP1 and ß-catenin. RIP1 expression can destroy the ß-catenin-ß-TrCP complex. Taken together, these results suggest a novel EMT-enhancing role of RIP1 in the WNT pathway and suggest a new canonical WNT3A-RIP1-ß-catenin pathway that contributes to CRC malignancy by promoting EMT.
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Neoplasias Colorretais , beta Catenina , Animais , Humanos , Camundongos , beta Catenina/genética , beta Catenina/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Neoplasias Colorretais/patologia , Regulação para Baixo , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Metástase Neoplásica , Via de Sinalização WntRESUMO
INTRODUCTION: The purpose of this study was to determine whether data preprocessing and augmentation could improve visual field (VF) prediction of recurrent neural network (RNN) with multi-central datasets. METHODS: This retrospective study collected data from five glaucoma services between June 2004 and January 2021. From an initial dataset of 331,691 VFs, we considered reliable VF tests with fixed intervals. Since the VF monitoring interval is very variable, we applied data augmentation using multiple sets of data for patients with more than eight VFs. We obtained 5,430 VFs from 463 patients and 13,747 VFs from 1,076 patients by setting the fixed test interval to 365 ± 60 days (D = 365) and 180 ± 60 days (D = 180), respectively. Five consecutive VFs were provided to the constructed RNN as input and the 6th VF was compared with the output of the RNN. The performance of the periodic RNN (D = 365) was compared to that of an aperiodic RNN. The performance of the RNN with 6 long- and short-term memory (LSTM) cells (D = 180) was compared with that of the RNN with 5-LSTM cells. To compare the prediction performance, the root mean square error (RMSE) and mean absolute error (MAE) of the total deviation value (TDV) were calculated as accuracy metrics. RESULTS: The performance of the periodic model (D = 365) improved significantly over aperiodic model. Overall prediction error (MAE) was 2.56 ± 0.46 dB versus 3.26 ± 0.41 dB (periodic vs. aperiodic) (p < 0.001). A higher perimetric frequency was better for predicting future VF. The overall prediction error (RMSE) was 3.15 ± 2.29 dB versus 3.42 ± 2.25 dB (D = 180 vs. D = 365). Increasing the number of input VFs improved the performance of VF prediction in D = 180 periodic model (3.15 ± 2.29 dB vs. 3.18 ± 2.34 dB, p < 0.001). The 6-LSTM in the D = 180 periodic model was more robust to worsening of VF reliability and disease severity. The prediction accuracy worsened as the false-negative rate increased and the mean deviation decreased. CONCLUSION: Data preprocessing with augmentation improved the VF prediction of the RNN model using multi-center datasets. The periodic RNN model predicted the future VF significantly better than the aperiodic RNN model.
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Pressão Intraocular , Campos Visuais , Humanos , Estudos Retrospectivos , Reprodutibilidade dos Testes , Testes de Campo Visual , Redes Neurais de Computação , Progressão da DoençaRESUMO
Phloretin is one of the apple polyphenols with anticancer activities. Since tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) serves important roles in inducing apoptosis, the present study examined the effect of phloretin on TRAIL-induced apoptosis in colon cancer cells. Treatment with both phloretin and TRAIL markedly suppressed the survival of cancer cells from several colon cancer cell lines compared with that of cells treated with either TRAIL or phloretin. Additionally, decreased numbers of colonies were observed following addition of phloretin and TRAIL. Furthermore, TRAIL- and phloretin-treated HT-29-Luc cells exhibited decreased luciferase activity. Increased apoptosis was observed in phloretin- and TRAIL-treated HT-29-Luc colon cancer cells, accompanying elevated levels of cleaved poly(ADP-ribose) polymerase, and caspase-3, -8 and -9. The expression levels of MCL1 apoptosis regulator BCL2 family member (Mcl-1) were decreased following addition of phloretin in colon cancer cells. In addition, overexpression of Mcl-1 in phloretin- and TRAIL-treated HT-29-Luc cells resulted in increased cell survival. Treatment of HT-29-Luc cells with a combination of cycloheximide (CHX) and phloretin led to a more prominent decrease in Mcl-1 expression compared with that in cells treated with CHX alone, while Mcl-1 expression was recovered by treatment with MG132. Binding of ubiquitin with Mcl-1 was verified using immunoprecipitation. Intraperitoneal injection of both TRAIL and phloretin into tumor xenografts was associated with a decreased tumor volume compared with that following injection with either TRAIL or phloretin. Overall, the present results suggest a synergistic effect of phloretin on TRAIL-induced apoptosis in colon cancer cells.
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Our team has previously reported a series of quinazoline-based lapatinib hybrids as potent kinase-targeting anticancer agents. Among them, AF8c showed a relatively safe profile in colorectal cancer (CRC) cells. In this study, we delineate a novel anticancer activity of AF8c in CRC cells. AF8c mediated p53-dependent apoptosis of CRC cells via the generation of endoplasmic reticulum (ER) stress and reactive oxygen species (ROS), as well as activation of nuclear respiratory factor 2 alpha subunit (Nrf2) and death receptor 5 (DR5), among others. The silencing of DR5 attenuated the expression levels of Nrf2 and partially inhibited AF8c-induced apoptosis. Additionally, upregulation of Nrf2 by AF8c evoked apoptosis through a decrease in antioxidant levels. Treatment of a CRC mice model with AF8c also resulted in the upregulation of DR5, Nrf2, and CHOP proteins, subsequently leading to a significant decrease in tumor burden. In comparison with lapatinib, AF8c showed higher cellular antiproliferative activity at the tested concentrations in CRC cells and synergized TRAIL effects in CRC cells. Overall, our results suggest that AF8c-induced apoptosis may be associated with DR5/Nrf2 activation through ER stress and ROS generation in CRC cells. These findings indicate that AF8c represents a promising polypharmacological molecule for the treatment of human CRC.
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BACKGROUND: Diagnosing Clostridioides difficile infection (CDI) is complicated. There have been reports on effects of compliance with anti-C. difficile prescription guidelines on patient outcomes. However, the causes of non-adherence and their impact on outcomes have rarely been explored. Therefore, an investigation on the risk factors for non-adherence with treatment guidelines and their influence on recurrence is important. METHODS: This case-control study was conducted with patients with a positive C. difficile culture from March 2020 to April 2021. We conducted analysis based on treatment categories using factors associated with recurrent CDI as variables. Univariate and multivariable analyses were conducted to identify risk factors for non-adherence with treatment guidelines. RESULTS: In total, culture positive stool samples from 172 patients were analyzed. Having positive glutamate dehydrogenase antigen (GDH Ag), negative toxin enzyme immunoassay (EIA), and positive nucleic acid amplification test (NAAT) (GDH+/toxin EIA-/NAAT +) results were associated with both under- (adjusted odds ratio [aOR] 3.49 [95% CI 1.62-7.51], p = 0.001) and over-treatment (aOR 0.17 [95% CI 0.06-0.48], p = 0.001). Patients with refractory diarrhea were over treated (aOR 2.71 [95% CI 1.02-7.20], p = 0.046). Patients with an increased risk of CDI recurrence were not over treated. CONCLUSIONS: Our results suggest that non-adherence with CDI treatment guidelines depends on the duration of symptoms and rapid EIA test results. Patients with an increased risk of recurrence were neglected.
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Toxinas Bacterianas , Clostridioides difficile , Infecções por Clostridium , Adulto , Antibacterianos/uso terapêutico , Estudos de Casos e Controles , Infecções por Clostridium/diagnóstico , Infecções por Clostridium/tratamento farmacológico , Humanos , SobretratamentoRESUMO
To assess the effect of Cannabidiol (CBD) on the angiogenesis and stemness of breast cancer cells as well as proliferation. Methods: mRNA level and the amount of protein of vascular endothelial growth factor (VEGF) were determined by qRT-PCR and ELISA. The angiogenic potential of breast cancer cells under hypoxic conditions was identified by the HUVEC tube formation assay. The degradation of HIF-1α by CBD and the Src/von Hippel-Lindau tumor suppressor protein (VHL) interaction were assessed by a co-immunoprecipitation assay and Western blotting. To identify the stemness of mamospheres, they were evaluated by the sphere-forming assay and flow cytometry. Results: CBD can suppress angiogenesis and stem cell-like properties of breast cancer through Src/VHL/HIF-1α signaling. CBD may potentially be utilized in the treatment of refractory or recurrent breast cancer.
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OBJECTIVES: This study was designed to investigate the molecular epidemiology of Clostridioides difficile isolates in South Korea and to evaluate risk factors for rifaximin-non-susceptible C. difficile infection (CDI). METHODS: A total of 413 patients with CDI from two sentinel hospitals in South Korea were enrolled in this study. Putative clinical risk factors for CDI were identified using digital medical records of the patients. Pathogen profiles, including antimicrobial susceptibility, toxin production and ribotype, were evaluated for each of the causative C. difficile isolates. RESULTS: Of the 413 C. difficile isolates, 81 (19.6%) were shown to be rifaximin-non-susceptible, with the most common ribotypes being 018 (56.8%; 46/81), 017 (16.0%; 13/81) and 027 (6.2%; 5/81). Rifaximin-non-susceptible C. difficile isolates exhibited higher non-susceptibility rates to most of the other drugs tested in this study compared with rifaximin-susceptible isolates. Previous history of pulmonary tuberculosis and prior rifaximin treatment were shown to be associated with the occurrence of rifaximin-non-susceptible CDI compared with susceptible CDI. CONCLUSION: Non-susceptibility rates to rifaximin for the C. difficile isolates identified in this study were reasonably high with most of the resistant strains belonging to either ribotype 018 or 017. Widespread dissemination of these clones may be the result of antimicrobial selection pressure introduced by the widespread use of rifaximin. These results suggest that a sustainable surveillance programme for CDI and C. difficile resistance is needed in order to better control CDIs and to improve therapeutic efficacy.
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Clostridioides difficile , Infecções por Clostridium , Clostridioides difficile/genética , Infecções por Clostridium/epidemiologia , Humanos , Testes de Sensibilidade Microbiana , Epidemiologia Molecular , Estudos Prospectivos , Rifaximina , Fatores de RiscoRESUMO
OBJECTIVES: Rapid detection of carbapenemase-producing Enterobacterales (CPE) is important to control spread of the resistance. We previously reported that imipenem disks prepared from injectable imipenem-cilastatin could rapidly detect KPC- and NDM-type carbapenemases. In the present study, we evaluated performance of disks of IPM and combined disks of imipenem-tazobactam and imipenem-EDTA, which were prepared from powders of imipenem and inhibitors. METHODS: Isolates of Enterobacterales were recovered from specimens of patients at a tertiary care hospital in Korea during January 2017 and March 2018. Routine CPE detection was performed by the CPE surveillance personnel whereas evaluation of the Disk carbapenemase test (DCT) was performed by the other personnel without knowing the results of surveillance. The DCT was carried out by pressing disks on to colonies and rehydrating in Petri plates and observing color change. RESULTS: The DCT differentiated 688 of 694 (sensitivity 99.1%) carbapenemase-producing isolates in 2.5-20 min: 630 with KPC, 51 with NDM, three with IMP, one with VIM, two with KPC and IMP, and one with NDM and OXA-181. The DCT failed to detect six OXA- 48-like enzyme-producing isolates, but the modified method using 96-well flat-bottom microplates with mineral oil cover detected all 29 OXA-48-like enzyme-producing isolates in 20-120 min. The DCT was negative for all 440 ertapenem-nonsusceptible, carbapenemase gene-negative isolates (specificity 100%). CONCLUSION: The procedure of DCT is simple and can differentiate isolates of Enterobacterales with KPC-, NDM-, IMP- and VIM-type carbapenemases rapidly, and the modified DCT can detect isolates with OXA-48-like enzymes rapidly.
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Infecções por Enterobacteriaceae , Proteínas de Bactérias , Humanos , República da Coreia , beta-LactamasesRESUMO
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is known to behave as an attractive anti-cancer agent in various cancers. Despite its promise TRAIL has limitations such as short half-life and rapid development of resistance. In this regard, approaches to sensitizers of TRAIL that can overcome the limitations of TRAIL are necessary. However, the molecular targets and mechanisms underlying sensitization to TRAIL-induced apoptosis are not fully understood. Here, we propose that reactive oxygen species modulator-1 (Romo1) as an attractive sensitizer of TRAIL. Romo1 is a mitochondrial inner membrane channel protein that controls reactive oxygen species (ROS) production, and its expression is highly upregulated in various cancers, including colorectal cancer. In the present study, we demonstrated that Romo1 inhibition significantly increased TRAIL-induced apoptosis of colorectal cancer cells, but not of normal colon cells. The combined effect of TRAIL and Romo1 inhibition was correlated with the activation of mitochondrial apoptosis pathways. Romo1 silencing elevated the protein levels of BCL-2-associated X protein (Bax) by downregulating the ubiquitin proteasome system (UPS). Romo1 inhibition downregulated the interaction between Bax and Parkin. Furthermore, Romo1 knockdown triggered the mitochondrial dysfunction and ROS generation. We validated the effect of combination in tumor xenograft model in vivo. In conclusion, our study demonstrates that Romo1 inhibition induces TRAIL-mediated apoptosis by identifying the novel mechanism associated with the Bax/Parkin interaction. We suggest that targeting of Romo1 is essential for the treatment of colorectal cancer and may be a new therapeutic approach in the future and contribute to the drug discovery.
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Oxaliplatin is used for treatment in combination with many drugs. However, the survival rate is still low due to side effects and drug resistance. Therefore, the combination with natural products was required for increasing efficacy and reducing side effects. Genipin, a natural product derived from the Gardenia jasminoides, associated with anti-angiogenic, anti-proliferative, hypertension, inflammatory, and the Hedgehog pathway. It is not known that genipin increases the therapeutic effect of oxaliplatin in gastric cancer. In this study, we found that genipin sensitizes oxaliplatin-induced apoptosis for the first time using colony forming assay, FACS analysis, and western blotting in gastric cancer. Additionally, genipin induced p53 expression in AGS, MKN45, and MKN28 cells. Also, genipin induced autophagy and LC3 expression. Knockdown of LC3 decreased cell death enhanced by the combination of oxaliplatin and genipin. In summary, we showed that genipin increases the oxaliplatin-induced cell death via p53-DRAM autophagy. Based on this, we suggest that genipin is a sensitizer of oxaliplatin.
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Disabled tumor suppressor genes and hyperactive oncogenes greatly contribute to cell fates during cancer development because of their genetic alterations such as somatic mutations. However, little is known about how tumor suppressor genes react to diverse oncogenes during tumor progression. Our previous study showed that RUNX3 inhibits invasiveness by preventing vascular endothelial growth factor secretion and suppressed endothelial cell growth and tube formation in colorectal cancer (CRC). Hedgehog signaling is crucial for the physiological maintenance and self-renewal of stem cells, and its deregulation is responsible for their tumor development. The mechanisms that inhibit this pathway during proliferation remain poorly understood. Here, we found that the tumor suppressor RUNX3 modulates tumorigenesis in response to cancer cells induced by inhibiting oncogene GLI1 ubiquitination. Moreover, we demonstrated that RUNX3 and GLI1 expression were inversely correlated in CRC cells and tissues. We observed a direct interaction between RUNX3 and GLI1, promoting ubiquitination of GLI1 at the intracellular level. Increased ubiquitination of GLI1 was induced by the E3 ligase ß-TrCP. This novel RUNX3-dependent regulatory loop may limit the extent and duration of Hedgehog signaling during extension of the tumor initiation capacity. On the basis of our results, identification of agents that induce RUNX3 may be useful for developing new and effective therapies for CRC.
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Neoplasias do Colo/metabolismo , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Proteínas Hedgehog/metabolismo , Animais , Apoptose , Ciclo Celular , Proliferação de Células , Neoplasias do Colo/patologia , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Transdução de Sinais , Células Tumorais CultivadasRESUMO
Hypoxia, or the deficiency of oxygen, in solid tumors is majorly responsible for the progression of cancer and remains unaffected by chemotherapy, but still requires definitive definition of the hypoxia signaling. Hypoxia disrupts the complete folding of mitochondrial proteins, leading to several diseases. The present study confirms that hypoxia activates the Hedgehog pathway in colorectal cancer (CRC), considering its role in cancer epithelial to mesenchymal transition, migration, and invasion. The activity of hypoxia-mediated Gli-1, a Hedgehog signaling factor in hypoxia, was confirmed by in vitro western blotting, immunofluorescence staining, wound-healing assay, and matrigel invasion assay, as well as by in vivo xenograft models (n = 5 per group). The Gli-1 mechanism in hypoxia was analyzed via mass spectrometry. Hypoxia enhanced the interaction of Gli-1 and T-complex protein 1 subunit beta (CCT2), as observed in the mass spectrometric analysis. We observed that reduction in CCT2 inhibits tumor induction by Gli-1. Ubiquitination-mediated Gli-1 degradation by ß-TrCP occurs during incomplete folding of Gli-1 in hypoxia. The human CRC tissues revealed greater CCT2 expression than did the normal colon tissues, indicating that higher CCT2 expression in tumor tissues from CRC patients reduced their survival rate. Moreover, we suggest that CCT2 correlates with Gli-1 expression and is an important determinant of survival in the CRC patients. The results reveal that CCT2 can regulate the folding of Gli-1 in relation to hypoxia in CRC.
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Chaperonina com TCP-1/genética , Neoplasias Colorretais/genética , Hipóxia Tumoral/genética , Proteína GLI1 em Dedos de Zinco/genética , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Chaperonina com TCP-1/química , Neoplasias Colorretais/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HCT116 , Proteínas Hedgehog/genética , Xenoenxertos , Humanos , Masculino , Camundongos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Dobramento de Proteína , Proteólise/efeitos dos fármacos , Transdução de Sinais/genética , Taxa de Sobrevida , Ubiquitinação/genética , Proteína GLI1 em Dedos de Zinco/química , Proteínas Contendo Repetições de beta-Transducina/farmacologiaRESUMO
According to recent studies, Cannabidiol (CBD), one of the main components of Cannabis sativa, has anticancer effects in several cancers. However, the exact mechanism of CBD action is not currently understood. Here, CBD promoted cell death in gastric cancer. We suggest that CBD induced apoptosis by suppressing X-linked inhibitor apoptosis (XIAP), a member of the IAP protein family. CBD reduced XIAP protein levels while increasing ubiquitination of XIAP. The expression of Smac, a known inhibitor of XIAP, was found to be elevated during CBD treatment. Moreover, CBD treatment increased the interaction between XIAP and Smac by increasing Smac release from mitochondria to the cytosol. CBD has also been shown to affect mitochondrial dysfunction. Taken together, these results suggest that CBD may have potential as a new therapeutic target in gastric cancer.
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Proteínas Reguladoras de Apoptose/metabolismo , Apoptose , Biomarcadores Tumorais/metabolismo , Canabidiol/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas Mitocondriais/metabolismo , Neoplasias Gástricas/patologia , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/metabolismo , Animais , Proteínas Reguladoras de Apoptose/genética , Biomarcadores Tumorais/genética , Proliferação de Células , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas Mitocondriais/genética , Invasividade Neoplásica , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Células Tumorais Cultivadas , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Despite the importance of radiation therapy, there are few radiation-related markers available for use in clinical practice. A larger catalog of such biomarkers is required to help clinicians decide when radiotherapy should be replaced with a patient-specific treatment. Arachidonate 15-lipoxygenase (15-LOX-1) enzyme is involved in polyunsaturated fatty acid metabolism. When colorectal cancer (CRC) cells were exposed to radiation, 15-LOX-1 was upregulated. To verify whether 15-LOX-1 protects against or induces DNA damage, we irradiated sh15-LOX-1 stable cells. We found that low 15-LOX-1 is correlated with radioresistance in CRC cells. These data suggest that the presence of 15-LOX-1 can be used as a marker for radiation-induced DNA damage. Consistent with this observation, gene-set-enrichment analysis based on microarray experiments showed that UV_RESPONSE was decreased in sh15-LOX-1 cells compared to shCon cells. Moreover, we discovered that the expression of the histone H2A variant macroH2A2 was sevenfold lower in sh15-LOX-1 cells. Overall, our findings present mechanistic evidence that macroH2A2 is transcriptionally regulated by 15-LOX-1 and suppresses the DNA damage response in irradiated cells by delaying H2AX activation.
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The molecular epidemiology and antimicrobial resistance of Clostridioides difficile were studied in South Korea in 2017 as part of a National Surveillance System. From February to May 2017, all non-duplicate isolates of C. difficile were recovered from patients who were suspected to have C. difficile infection and collected from 6 referral hospitals representing the 6 regions in South Korea. We performed PCRs for the toxin gene, PCR ribotyping, multilocus sequence typing (MLST), antimicrobial susceptibility testing by agar dilution according to the recommendations of the CLSI and detection of antimicrobial resistance genes such as ermB, catD, tetM, vanZ and nimR by PCR. Of 331 C. difficile isolates, 257 (77.6%) were toxigenic and the prevalence of strains producing binary toxin (CDT) was 5.1% (13/257). A total of 52 different ribotype (RT) patterns were found. RT018 was the most common (25.1% of all isolates), and RT014/020, RT002 and RT012 were also common. RT010 was most common non-toxigenic strain. MLST analysis of randomly selected 72 C. difficile isolates identified 46 sequence types (STs), of which three were new and not in the PubMLST library. There was a good correlation between MLST and RT as following: ST1 (RT027), ST8 (RT002), ST11 (RT078), ST17 (RT018), ST35 (RT046), ST37 (RT017), ST42 (RT106), ST53 (RT103), ST81 (RT369), and ST99 (RT070). All toxigenic isolates were susceptible to metronidazole and vancomycin (MICâ¯≤â¯2â¯mg/L). For rifaximin, 24% of toxigenic isolates were resistant. Of randomly selected 106 toxigenic isolates, resistance rates for ampicillin, cefotetan, clindamycin, imipenem, chloramphenicol, tetracycline, and moxifloxacin were 48%, 46%, 64%, 54%, 0%, 6% and 52% respectively and frequencies of various resistance genes were 62.3% for ermB, 0.9% catD and 10.4% tetM. RTs018, 002, 017 and 369 showed high MICs to various antimicrobial agents and multi-drug resistance was common also.
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Antibacterianos/farmacologia , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/genética , Infecções por Clostridium/epidemiologia , Infecções por Clostridium/microbiologia , Adulto , Idoso , Antibacterianos/uso terapêutico , Toxinas Bacterianas/genética , Clostridioides difficile/classificação , Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/tratamento farmacológico , Infecções Comunitárias Adquiridas/epidemiologia , Infecções Comunitárias Adquiridas/microbiologia , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , Farmacorresistência Bacteriana , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Epidemiologia Molecular , Tipagem de Sequências Multilocus , Vigilância em Saúde Pública , República da Coreia/epidemiologia , RibotipagemRESUMO
BACKGROUND: Although the anticancer activity of Korean Red Ginseng (KRG) has been known in various cancers, the mechanism of KRG-induced apoptosis is unknown in colorectal cancer (CRC). In our study, we examined whether KRG induces apoptosis in CRC cells. METHODS: In the cell viability assay, the concentration of the appropriate KRG extracts was fixed at 2.5 mg/mL in numerous CRC cells. This fixed concentration was in other experiments, and it was confirmed that the KRG extracts induce apoptosis in CRC cells. RESULTS: We found that KRG induced Noxa activation and apoptosis and increased endoplasmic reticulum stress via reactive oxygen species production. This indicated that KRG efficiently enhanced cell death in CRC cells. CONCLUSION: Our results show that KRG can be used as a possible anticancer drug for patients with CRC.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias Colorretais/tratamento farmacológico , Panax/química , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Antineoplásicos Fitogênicos/isolamento & purificação , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Células HT29 , Humanos , Extratos Vegetais/isolamento & purificação , Proteínas Proto-Oncogênicas c-bcl-2/genética , Espécies Reativas de Oxigênio/metabolismo , Transdução de SinaisRESUMO
BACKGROUND: Genipin is a compound derived from gardenia fruit extract. Although Genipin has anti-tumor effects in various cancers, its effect and mechanism in gastric cancer remain unclear. Here, we investigated the relationship between the anticancer effect of Genipin and signal transducer and activator of transcription (Stat3)/myeloid cell leukemia-1 (Mcl-1) in human gastric cancers. METHODS: MTT assays were performed to determine the cell viability of gastric cancer and gastric epithelial cell lines (AGS, MKN45, SNU638, MKN74, HFE-145). A TUNEL assay and Western blotting were carried out to investigate apoptosis. Stat3 activity was measured by proteome profiler phospho kinase array, immunofluorescence and immunoblotting. Mitochondria function was monitored with an XF24 analyzer and by flow cytometry, confocal microscopy using fluorescent probes for general mitochondrial membrane potential (MMP). RESULTS: Genipin induced apoptosis in gastric cancer cells, including AGS and MKN45 cells. Genipin also reduced Mcl-1 mRNA and protein levels. Furthermore, we found that phosphorylation of Stat3 is regulated by Genipin. Additionally, the protein level of phospho Janus kinase 2 (JAK2) was decreased by Genipin treatment, indicating that the Stat3/JAK2/Mcl-1 pathway is suppressed by Genipin treatment in gastric cancer cells. Mcl-1 is closely related to mitochondrial function. These findings suggest that Genipin contributes to the collapse of mitochondrial functions like MMP. CONCLUSIONS: Genipin induced apoptosis by suppressing the Stat3/Mcl-1 pathway and led to mitochondrial dysfunction. Our results reveal a novel mechanism for the anti-cancer effect of Genipin in gastric cancer.