RESUMO
As people age, the risk and progression of colorectal cancer (CRC), along with cholesterol levels, tend to increase. Nevertheless, epidemiological studies on serum lipids and CRC have produced conflicting results. We previously demonstrated that the reduction of squalene epoxidase (SQLE) due to accumulated cholesterol within cells accelerates CRC progression through the activation of the ß-catenin pathway. This study aimed to investigate the mechanism by which age-related cholesterol accumulation within tissue accelerates CRC progression and to assess the clinical significance of SQLE in older individuals with elevated CRC risk. Using machine learning-based digital image analysis with fluorescence-immunohistochemistry, we assessed SQLE, GSK3ßpS9 (GSK3ß activity inhibition through serine 9 phosphorylation at GSK3ß), p53 wild-type (p53WT), and p53 mutant (p53MT) levels in CRC tissues. Our analysis revealed a significant reduction in SQLE, p53WT, and p53MT and increase in GSK3ßpS9 levels, all associated with the substantial accumulation of intra-tissue cholesterol in aged CRCs. Cox analysis underscored the significant influence of SQLE on overall survival and progression-free survival in grade 2-3 CRC patients aged over 50. SQLE and GSK3ßpS9 consistently exhibited outstanding prognostic and diagnostic performance, particularly in older individuals. Furthermore, combining SQLE with p53WT, p53MT, and GSK3ßpS9 demonstrated a robust diagnostic ability in the older population. In conclusion, we have identified that individuals aged over 50 face an increased risk of CRC progression due to aging-linked cholesterol accumulation within tissue and the subsequent reduction in SQLE levels. This study also provides valuable biomarkers, including SQLE and GSK3ßpS9, for older patients at elevated risk of CRC.
Assuntos
Colesterol , Neoplasias Colorretais , Progressão da Doença , Esqualeno Mono-Oxigenase , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Envelhecimento/metabolismo , Colesterol/metabolismo , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/genética , Fatores de Risco , Esqualeno Mono-Oxigenase/metabolismo , Esqualeno Mono-Oxigenase/genéticaRESUMO
Recently, we reported the involvement of TIPRL/LC3/CD133 in liver cancer aggressiveness. This study assessed the human TOR signaling regulator (TIPRL)/microtubule-associated light chain 3 (LC3)/prominin-1 (CD133)/cluster of differentiation 44 (CD44) as potential diagnostic and prognostic biomarkers for early liver cancer. For the assessment, we stained tissues of human liver disease/cancer with antibodies against TIPRL/LC3/CD133/CD44/CD46, followed by confocal observation. The roles of TIPRL/LC3/CD133/CD44/CD46 in liver normal and cancer cell lines were determined by in vitro studies. We analyzed the prognostic and diagnostic potentials of TIPRL/LC3/CD133/CD44/CD46 using the receiver-operating characteristic curve, a Kaplan-Meier and uni-/multi-Cox analyses. TIPRL and LC3 were upregulated in tissues of HCCs and adult hepatocytes-derived liver diseases while downregulated in iCCA. Intriguingly, TIPRL levels were found to be critically associated with liver cancer patients' survivability, and TIPRL is the key player in liver cancer cell proliferation and viability via stemness and self-renewal induction. Furthermore, we demonstrate that TIPRL/LC3/CD133 have shown prominent efficiency for diagnosing patients with grade 1 iCCA. TIPRL/LC3/CD133/CD44 have also provided excellent potential for prognosticating patients with grade 1 iCCA and grade 1 HCCs, together with demonstrating that TIPRL/LC3/CD133/CD44 are, either individually or in conjunction, potential biomarkers for early liver cancer.
RESUMO
BACKGROUND & AIMS: Squalene epoxidase (SQLE), a rate-limiting enzyme in cholesterol biosynthesis, is suggested as a proto-oncogene. Paradoxically, SQLE is degraded by excess cholesterol, and low SQLE is associated with aggressive colorectal cancer (CRC). Therefore, we studied the functional consequences of SQLE reduction in CRC progression. METHODS: Gene and protein expression data and clinical features of CRCs were obtained from public databases and 293 human tissues, analyzed by immunohistochemistry. In vitro studies showed underlying mechanisms of CRC progression mediated by SQLE reduction. Mice were fed a 2% high-cholesterol or a control diet before and after cecum implantation of SQLE genetic knockdown/control CRC cells. Metastatic dissemination and circulating cancer stem cells were demonstrated by in vivo tracking and flow cytometry analysis, respectively. RESULTS: In vitro studies showed that SQLE reduction helped cancer cells overcome constraints by inducing the epithelial-mesenchymal transition required to generate cancer stem cells. Surprisingly, SQLE interacted with GSK3ß and p53. Active GSK3ß contributes to the stability of SQLE, thereby increasing cell cholesterol content, whereas SQLE depletion disrupted the GSK3ß/p53 complex, resulting in a metastatic phenotype. This was confirmed in a spontaneous CRC metastasis mice model, where SQLE reduction, by a high-cholesterol regimen or genetic knockdown, strikingly promoted CRC aggressiveness through the production of migratory cancer stem cells. CONCLUSIONS: We showed that SQLE reduction caused by cholesterol accumulation aggravates CRC progression via the activation of the ß-catenin oncogenic pathway and deactivation of the p53 tumor suppressor pathway. Our findings provide new insights into the link between cholesterol and CRC, identifying SQLE as a key regulator in CRC aggressiveness and a prognostic biomarker.
Assuntos
Colesterol/metabolismo , Neoplasias Colorretais/patologia , Esqualeno Mono-Oxigenase/metabolismo , Adulto , Idoso , Animais , Linhagem Celular Tumoral , Colo/patologia , Modelos Animais de Doenças , Feminino , Técnicas de Silenciamento de Genes , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Mucosa Intestinal/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/patologia , Oxirredução , Proto-Oncogene Mas , Reto/patologia , Esqualeno Mono-Oxigenase/genética , Proteína Supressora de Tumor p53/metabolismo , Adulto Jovem , beta Catenina/metabolismoRESUMO
The coiled-coil domain containing 50 (CCDC50) protein is a phosphotyrosine-dependent signalling protein stimulated by epidermal growth factor. It is highly expressed in neuronal cells in the central nervous system; however, the roles of CCDC50 in neuronal development are largely unknown. In this study, we showed that the depletion of CCDC50-V2 impeded the neuronal development process, including arbor formation, spine density development, and axonal outgrowth, in primary neurons. Mechanistic studies revealed that CCDC50-V2 positively regulated the nerve growth factor receptor, while it downregulated the epidermal growth factor receptor pathway. Importantly, JNK/c-Jun activation was found to be induced by the CCDC50-V2 overexpression, in which the interaction between CCDC50-V2 and JNK2 was also observed. Overall, the present study demonstrates a novel mechanism of CCDC50 function in neuronal development and provides new insight into the link between CCDC50 function and the aetiology of neurological disorders.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Crescimento Neuronal , Animais , Linhagem Celular Tumoral , Receptores ErbB/metabolismo , Células HEK293 , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Camundongos , Proteínas do Tecido Nervoso/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Receptores de Fator de Crescimento Neural/metabolismo , Transdução de SinaisRESUMO
Autophagy, an intracellular system of degrading damaged organelles and misfolded proteins, is essential for cancer cell survival. Despite the progress made towards understanding the mechanism, identification of novel autophagy regulators presents a major obstacle in developing anticancer therapies. Here, we examine the association between the TOR signaling pathway regulator-like (TIPRL) protein and autophagy in malignant transformation of tumors. We show that TIPRL upregulation in non-small cell lung cancer (NSCLC) potentiated autophagy activity and enabled autophagic clearance of metabolic and cellular stress, conferring a survival advantage to cancer cells. Importantly, the interaction of TIPRL with eukaryotic initiation factor 2α (eIF2α) led to eIF2α phosphorylation and activation of the eIF2α-ATF4 pathway, thereby inducing autophagy. Conversely, TIPRL depletion increased apoptosis by reducing autophagic clearance, which was markedly enhanced in TIPRL-depleted A549 xenografts treated with 2-deoxy-D-glucose. Overall, the study indicated that TIPRL is a potential regulator of autophagy and an important drug target for lung cancer therapy.
Assuntos
Fator 4 Ativador da Transcrição/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Fator de Iniciação 2 em Eucariotos/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Pulmonares/genética , Células A549 , Animais , Apoptose , Autofagia/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Sobrevivência Celular , Estresse do Retículo Endoplasmático , Feminino , Xenoenxertos , Humanos , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Fosforilação , Transdução de Sinais , Esferoides Celulares/patologiaRESUMO
Studies have reported dysregulation of TIPRL, LC3 and CD133 in liver cancer tissues. However, their respective relationships to liver cancer and roles as biomarkers for prognosis and diagnosis of liver cancer have never been studied. Here we report that the level of TIPRL is significantly correlated with levels of LC3 (Spearman r = 0.9) and CD133 (r = 0.7) in liver cancer tissues. We observed significant upregulations of TIPRL, LC3 and CD133 in hepatocellular carcinomas (HCCs) compared with adjacent normal tissues. Importantly, TIPRL, tested among additional variables, showed a significant impact on the prognosis of HCC patients. TIPRL knockdown significantly reduced expressions of LC3, CD133, stemness-related genes, as well as viability and stemness of liver cancer cell-lines, which were promoted by ectopic TIPRL expression. Either alone or as a combination, TIPRL, LC3 and CD133 showed significant values of area under the curve (AUC) and sensitivity/specificity in early liver cancer tissues. Furthermore, the statistical association and the diagnostic efficacies of TIPRL, LC3 and CD133 in HCC tissues were confirmed in a different IHC cohort. This data demonstrates that the complex involvement of TIPRL/LC3/CD133 in liver cancer aggressiveness can together or individually serve as potential biomarkers for the early detection of liver cancer.
Assuntos
Antígeno AC133/metabolismo , Carcinoma Hepatocelular/diagnóstico , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Hepáticas/diagnóstico , Proteínas Associadas aos Microtúbulos/metabolismo , Regulação para Cima , Antígeno AC133/genética , Área Sob a Curva , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Estudos de Coortes , Detecção Precoce de Câncer , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Proteínas Associadas aos Microtúbulos/genética , Células-Tronco Neoplásicas/metabolismo , PrognósticoRESUMO
Hepatocellular carcinoma (HCC) is one of the most malignant tumors. Although various treatments, such as surgery and chemotherapy, have been developed, a novel alternative therapeutic approach for HCC therapy is urgently needed. Tumor necrosis factor-related apoptosis inducing ligand (TRAIL) is a promising anti-cancer agent, but many cancer cells are resistant to TRAIL-induced apoptosis. To help overcome TRAIL resistance in HCC cancer cells, we have identified novel chemical compounds that act as TRAIL sensitizers. We first identified the hit compound, TRT-0002, from a chemical library of 6,000 compounds using a previously developed high-throughput enzyme-linked immunosorbent assay (ELISA) screening system, which was based on the interaction of mitogen-activated protein kinase kinase 7 (MKK7) and TOR signaling pathway regulator-like (TIPRL) proteins and a cell viability assay. To increase the efficacy of this TRAIL sensitizer, we synthesized 280 analogs of TRT-0002 and finally identified two lead compounds (TRT-0029 and TRT-0173). Co-treating cultured Huh7 cells with either TRT-0029 or TRT-0173 and TRAIL resulted in TRAIL-induced apoptosis due to the inhibition of the MKK7-TIPRL interaction and subsequent phosphorylation of MKK7 and c-Jun N-terminal kinase (JNK). In vivo, injection of these compounds and TRAIL into HCC xenograft tumors resulted in tumor regression. Taken together, our results suggest that the identified lead compounds serve as TRAIL sensitizers and represent a novel strategy to overcome TRAIL resistance in HCC.
RESUMO
Tumor metastasis is the leading cause of cancer death. In the metastatic process, EMT is a unique phenotypic change that plays an important role in cell invasion and changes in cell morphology. Despite the clinical significance, the mechanism underlying tumor metastasis is still poorly understood. Here we report a novel mechanism by which secreted plasma glutamate carboxypeptidase(PGCP) negatively involves Wnt/ß-catenin signaling by DKK4 regulation in liver cancer metastasis. Pathway analysis of the RNA sequencing data showed that PGCP knockdown in liver cancer cell lines enriched the functions of cell migration, motility and mesenchymal cell differentiation. Depletion of PGCP promoted cell migration and invasion via activation of Wnt/ß-catenin signaling pathway components such as phospho-LRP6 and ß-catenin. Also, addition of DKK4 antagonized the Wnt/ß-catenin signaling cascade in a thyroxine (T4)-dependent manner. In an in vivo study, metastatic nodules were observed in the lungs of the mice after injection of shPGCP stable cell lines. Our findings suggest that PGCP negatively associates with Wnt/ß-catenin signaling during metastasis. Targeting this regulation may represent a novel and effective therapeutic option for liver cancer by preventing metastatic activity of primary tumor cells.
Assuntos
Carboxipeptidases/sangue , Movimento Celular , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/patologia , Animais , Carboxipeptidases/antagonistas & inibidores , Carboxipeptidases/genética , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Feminino , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Hepáticas/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Metástase Neoplásica , RNA Interferente Pequeno/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Via de Sinalização Wnt/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
TRAIL (TNF-related apoptosis inducing ligand) is a promising anti-cancer drug target that selectively induces apoptosis in cancer cells. However, many cancer cells are resistant to TRAIL-induced apoptosis. Therefore, reversing TRAIL resistance is an important step for the development of effective TRAIL-based anti-cancer therapies. We previously reported that knockdown of the TOR signaling pathway regulator-like (TIPRL) protein caused TRAIL-induced apoptosis by activation of the MKK7-c-Jun N-terminal Kinase (JNK) pathway through disruption of the MKK7-TIPRL interaction. Here, we identified Taraxacum officinale F.H. Wigg (TO) as a novel TRAIL sensitizer from a set of 500 natural products using an ELISA system and validated its activity by GST pull-down analysis. Furthermore, combination treatment of Huh7 cells with TRAIL and TO resulted in TRAIL-induced apoptosis mediated through inhibition of the MKK7-TIPRL interaction and subsequent activation of MKK7-JNK phosphorylation. Interestingly, HPLC analysis identified chicoric acid as a major component of the TO extract, and combination treatment with chicoric acid and TRAIL induced TRAIL-induced cell apoptosis via JNK activation due to inhibition of the MKK7-TIPRL interaction. Our results suggest that TO plays an important role in TRAIL-induced apoptosis, and further functional studies are warranted to confirm the importance of TO as a novel TRAIL sensitizer for cancer therapy. © 2015 Wiley Periodicals, Inc.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Hepáticas/tratamento farmacológico , Ligante Indutor de Apoptose Relacionado a TNF/farmacologia , Taraxacum/química , Antineoplásicos Fitogênicos/química , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Neoplasias Hepáticas/metabolismo , MAP Quinase Quinase 7/metabolismo , Mapas de Interação de Proteínas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Ligante Indutor de Apoptose Relacionado a TNF/metabolismoRESUMO
Induction of apoptosis through activation of the TRAIL pathway is considered to be a promising anticancer strategy due to its ability to selectively induce apoptosis in cancer cells. However, the ability of cancer cells to acquire TRAIL resistance has limited the clinical translation of this approach. We previously reported that the TOR signaling pathway regulator-like (TIPRL) protein contributes to the resistance to TRAIL-induced apoptosis by inhibiting the MKK7-c-Jun N-terminal kinase (JNK) pathway via MKK7TIPRL interaction. In the present study, we identified Tussilago farfara L. (TF) as a novel TRAIL sensitizer among 500 natural products using an ELISA system that specifically detects the MKK7-TIPRL interaction, and we validated candidates by GST-pull down assay. Co-treatment of Huh7 cells with TF and TRAIL induced apoptosis via inhibition of the MKK7-TIPRL interaction and an increase in MKK7/JNK phosphorylation. This is the first report to describe TF as a novel TRAIL sensitizer, unveiling a potentially novel therapeutic strategy in cancer therapy.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Carcinoma Hepatocelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Hepáticas/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Extratos Vegetais/farmacologia , Tussilago/química , Apoptose , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologiaRESUMO
When an inflammatory stimulus is given, vascular endothelial cells express various cell adhesion molecules including the vascular cell adhesion molecule (VCAM)-1. In this study, the possibility of specifically delivering anti-inflammatory drugs to activated endothelial cells by utilizing VCAM-1 as a target receptor was explored by loading celecoxib, a selective cyclooxygenase-2 inhibitor, into liposomes coupled to the Fab' fragment against VCAM-1. Anti-VCAM-1-Fab'-conjugated liposomes were prepared by forming an amide linkage between amino groups of Fab' and the carboxylic group of glutaryl-N-phosphatidylethanolamine in liposomes using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide as a cross-linker in the presence of sulpho-N-hydroxysuccinimide. The coupling of Fab' to phospholipids constituting liposomes was confirmed by SDS-PAGE analysis. Under our optimized conjugation conditions, 130.0 µg Fab' was coupled to 1 µmol liposomes. Immunoblotting analysis showed that VCAM-1 protein expression could be induced by incubating human umbilical vein endothelial cells (HUVEC) with TNF-α. Confocal laser microsopy analysis revealed that Fab' conjugation to liposomes selectively increased liposomal uptake in TNF-α-pre-stimulated (VCAM-1-expressed) HUVECs, but not in cells without VCAM-1 expression. The concentration of celecoxib loaded in Fab'-conjugated liposomes was 281.1 ± 29 µg/mL, suggesting that liposomal loading also helped to overcome the limitations in celecoxib administration caused by its poor water solubility. Celecoxib loaded in Fab'-conjugated liposomes inhibited prostaglandin E2 (PGE2) production induced by TNF-α-pre-stimulation more efficiently than when loaded in conventional liposomes. Therefore, Fab'-conjugated liposomes served as a drug delivery system with dual functions: targeted delivery and solubilizing capacity.
Assuntos
Células Endoteliais/citologia , Fragmentos Fab das Imunoglobulinas/química , Pirazóis , Sulfonamidas , Veias Umbilicais/citologia , Molécula 1 de Adesão de Célula Vascular/química , Celecoxib , Células Cultivadas , Inibidores de Ciclo-Oxigenase 2/química , Inibidores de Ciclo-Oxigenase 2/farmacocinética , Inibidores de Ciclo-Oxigenase 2/farmacologia , Células Endoteliais/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Lipossomos , Pirazóis/química , Pirazóis/farmacocinética , Pirazóis/farmacologia , Solubilidade , Sulfonamidas/química , Sulfonamidas/farmacocinética , Sulfonamidas/farmacologia , Fator de Necrose Tumoral alfa/biossíntese , Veias Umbilicais/metabolismo , Molécula 1 de Adesão de Célula Vascular/biossíntese , Molécula 1 de Adesão de Célula Vascular/imunologia , Água/químicaRESUMO
PURPOSE: A retrorectal developmental cyst (tailgut cyst, epidermoid cyst, dermoid cyst, teratoma, and duplication) is very rare disease, and the symptoms are not characteristic so that sometimes this disease is still misdiagnosed as a supralevator abscess or a complex anal fistula. We would like to present a clinical approach to this disease. METHODS: We retrospectively examined the charts of 15 patients who were treated for retrorectal cysts from January 2001 to November 2009. RESULTS: All 15 patients were female. The average age was 41 years (range, 21 to 60 years). Fourteen patients (93.3%) were symptomatic, and the most common symptom was anal pain or discomfort. Nine patients (60%) had more than one previous operation (range, 1 to 9 times) for a supralevator abscess, an anal fistula, etc. In 12 patients (80%), the diagnosis could be made by using the medical history and physical examination. Thirteen cysts (80%) were excised completely through the posterior approach. The average diameter of the cysts was 4.8 cm (range, 2 to 10 cm). Pathologic diagnoses were 8 tailgut cysts (53.3%), 5 epidermoid cysts (33.3%) and 2 dermoid cysts (13.3%). The average follow-up period was 18.3 months (range, 1 to 64 months). CONCLUSION: In our experience, high suspicion and physical examination are the most important diagnostic methods. If a female patient has a history of multiple perianal operations, a retrorectal bulging soft mass, a posterior anal dimple, and no conventional creamy foul odorous pus in drainage, the possibility of a retrorectal developmental cyst must be considered.