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Predictive biomarkers of response to chemotherapy in patients with metastatic colorectal cancer (mCRC) are needed to better characterize tumors and enable more tailored therapies. Here we used serum proteomics to screen for chemotherapy predictive markers. We found that higher baseline serum inter-α-trypsin inhibitor Heavy Chain 4 (ITIH4) expression in newly diagnosed mCRC patients was associated with poorer response to standard first-line chemotherapy. In addition, the higher expression of ITIH4 in CRC tissue also suggested poorer prognosis mCRC patients. Moreover, the overexpression of ITIH4 could promote the proliferation of CRC cells and reduce the sensitivity of CRC cells to 5-fluorouracil (5-FU) by inhibiting apoptosis in vivo and vitro. Through RNA-seq combined with bioinformatics analysis, we speculated that ITIH4 may activate phosphatidyl 3-kinase-protein kinase B (PI3K-AKT) pathway to inhibit apoptosis, thereby reducing the sensitivity of CRC cells to 5-FU. In conclusion, our findings unveil that ITIH4 is associated with CRC resistance to 5-FU, and may serve as a potential predictive biomarker for the sensitivity of advanced CRC patients to standard first-line chemotherapy regimens, and also provide a potential therapeutic target to render 5-FU resistance in CRC patients.
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Surface-assisted laser desorption/ionization (SALDI) mass spectrometry imaging (MSI) holds great value in spatial metabolomics and tumor diagnosis. Tissue imprinting on the SALDI target can avoid laser-induced tissue ablation and simplifies the sample preparation. However, the tissue imprinting process always causes lateral diffusion of biomolecules, thereby losing the fidelity of metabolite distribution on tissue. Herein, a membrane-mediated imprinting mass spectrometry imaging (MMI-MSI) strategy is proposed using isoporous nuclepore track-etched membrane as a mediating imprinting layer to selectively transport metabolites through uniform and vertical pores onto silicon nanowires (SiNWs) array. Compared with conventional direct imprinting technique, MMI-MSI can not only exclude the adsorption of large biomolecules but also avoid the lateral diffusion of metabolites. The whole time for MMI-based sample preparation can be reduced to 2 min, and the lipid peak number can increase from 46 to 113 in kidney tissue detection. Meanwhile, higher resolution of MSI can be achieved due to the confinement effect of the pore channel in the diffusion of metabolites. Based on MMI-MSI, the tumor margins of liver cancer can be clearly discriminated and their different subtypes can be precisely classified. This work demonstrates MMI-MSI is a rapid, highly sensitive, robust and high-resolution technique for spatially-resolved metabolomics and pathological diagnosis.
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Efficient peptide and protein identifications from data-independent acquisition mass spectrometric (DIA-MS) data typically rely on a project-specific spectral library with a suitable size. Here, we describe subLib, a computational strategy for optimizing the spectral library for a specific DIA data set based on a comprehensive spectral library, requiring the preliminary analysis of the DIA data set. Compared with the pan-human library strategy, subLib achieved a 41.2% increase in peptide precursor identifications and a 35.6% increase in protein group identifications in a test data set of six colorectal tumor samples. We also applied this strategy to 389 carcinoma samples from 15 tumor data sets: up to a 39.2% increase in peptide precursor identifications and a 19.0% increase in protein group identifications were observed. Our strategy for spectral library size optimization thus successfully proved to deepen the proteome coverages of DIA-MS data.
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Neoplasias , Proteoma , Humanos , Espectrometría de Masas , Biblioteca de Péptidos , Péptidos/análisis , Proteoma/análisis , Proteómica/métodosRESUMEN
Lung cancer (LC) is a widespread cancer that is the cause of the highest mortality rate accounting for 25% of all cancer deaths. To date, most LC patients are diagnosed at the advanced stage owing to the lack of obvious symptoms in the early stage and the limitations of current clinical diagnostic techniques. Therefore, developing a high throughput technique for early screening is of great importance. In this work, we established an effective and rapid salivary metabolic analysis platform for early LC diagnosis and combined metabolomics and transcriptomics to reveal the metabolic fluctuations correlated to LC. Saliva samples were collected from a total of 150 volunteers including 89 patients with early LC, 11 patients with advanced LC, and 50 healthy controls. The metabolic profiling of noninvasive samples was investigated on an ultralow noise TELDI-MS platform. In addition, data normalization methods were screened and assessed to overcome the MS signal variation caused by individual difference for biomarker mining. For untargeted metabolic profiling of saliva samples, around 264 peaks could be reliably detected in each sample. After multivariate analysis, 23 metabolites were sorted out and verified to be related to the dysfunction of the amino acid and nucleotide metabolism in early LC. Notably, transcriptomic data from online TCGA repository were utilized to support findings from the salivary metabolomics experiment, including the disorder of amino acid biosynthesis and amino acid metabolism. Based on the verified differential metabolites, early LC patients could be clearly distinguished from healthy controls with a sensitivity of 97.2% and a specificity of 92%. The ultralow noise TELDI-MS platform displayed satisfactory ability to explore salivary metabolite information and discover potential biomarkers that may help develop a noninvasive screening tool for early LC.
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Neoplasias Pulmonares , Saliva , Humanos , Rayos Láser , Neoplasias Pulmonares/diagnóstico , Espectrometría de Masas , MetabolómicaRESUMEN
BACKGROUND: The role and mechanism of the nicotinamide adenine dinucleotide (NAD+) salvage pathway in cancer cell proliferation is poorly understood. Nicotinamide phosphoribosyltransferase (NAMPT), which converts nicotinamide into NAD+, is the rate-limiting enzyme in the NAD+ salvage pathway. Here, we assessed the role of NAMPT in the proliferation of colorectal cancer. METHODS: Real-time PCR, immunohistochemistry, western blotting, and analyses of datasets from Oncomine and Gene Expression Omnibus were conducted to assess the expression of NAMPT at the mRNA and protein levels in colorectal cancer. The Kaplan Meier plotter online tool was used to evaluate the prognostic role of NAMPT. Knockdown of NAMPT was performed to assess the role of NAMPT in colorectal cancer cell proliferation and tumorigenesis both in vitro and in vivo. Overexpression of NAMPT was used to evaluate impact of NAMPT on colorectal cancer cell proliferation in vitro. NAD+ quantitation, immunofluorescence, dual luciferase assay and western blot were used to explore the mechanism of colorectal cancer proliferation. Transwell migration and invasion assays were conducted to assess the role of NAMPT in cell migration and invasion abilities of colorectal cancer cells. RESULTS: Our study indicated that the inhibition of NAMPT decreased proliferation capacity of colorectal cancer cells both in vitro and in vivo. Conversely, overexpression of NAMPT could promote cell proliferation in vitro. NAMPT inhibition induced ß-catenin degradation by increasing Axin expression levels; this resulted in the inhibition of Wnt/ß-catenin signaling and cell proliferation in colorectal cancer. The addition of nicotinamide mononucleotide, the enzymatic product of NAMPT, effectively reversed ß-catenin protein degradation and inhibited growth. Similarly, the knockdown of Axin also decreased the cell death induced by the inhibition of NAMPT. In addition, we showed that colorectal cancer tissues harbored significantly higher levels of NAMPT than the levels harbored by paired normal tissues, especially in colorectal cancer stages I and II. And the overexpression of NAMPT was associated with unfavorable survival results. CONCLUSIONS: Our findings reveal that NAMPT plays an important role in colorectal cancer proliferation via Wnt/ß-catenin pathway, which could have vital implications for the diagnosis, prognosis and treatment of colorectal cancer.
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Proteína Axina/metabolismo , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Mutación/genética , NAD/metabolismo , Vía de Señalización Wnt , Acrilamidas/farmacología , Proteína de la Poliposis Adenomatosa del Colon/genética , Animales , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Citocinas/antagonistas & inhibidores , Citocinas/metabolismo , Femenino , Técnicas de Silenciamiento del Gen , Humanos , Ratones Endogámicos BALB C , Ratones Desnudos , Modelos Biológicos , Invasividad Neoplásica , Nicotinamida Fosforribosiltransferasa/antagonistas & inhibidores , Nicotinamida Fosforribosiltransferasa/metabolismo , Piperidinas/farmacología , Pronóstico , Vía de Señalización Wnt/efectos de los fármacosRESUMEN
Oxidative RNA damage has been found to be associated with a variety of diseases, and 8-hydroxyguanosine (8-OHG) is a typical marker of oxidative modification of RNA. This guanosine modification is an emerging biomarker for disease detection and determination of 8-OHG in human urine is favored because it is noninvasive to patients. However, due to its poor ionization efficiency in mass spectrometry and trace amount in urine, accurate quantification of this modified nucleoside is still challenging. Herein, a rapid, accurate, sensitive and robust method using solid-phase extraction (SPE) combined with isotope dilution ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) was developed for detection of this oxidative RNA modification in human urine. The limit of detection can reach 1.5 fmol and the method exhibits good precision on intra-day (1.8-3.3%) and inter-day (0.6-1.2%) analyses. Satisfactory recovery (87.5-107.2%) at three spiked levels was achieved by using HLB cartridge for urine pretreatment. Using this method, we quantified 8-OHG in urine from 65 colorectal cancer (CRC) patients and 76 healthy volunteers. The measured level of urinary 8-OHG for CRC patients and healthy controls is 1.91 ± 0.63 nmol/mmol creatinine and 1.33 ± 0.35 nmol/mmol creatinine, respectively. We found the content of 8-OHG in urine was raised in CRC patients patients, implying this oxidative RNA modification marker could act as a potential noninvasive indicator for early screening of CRC. In addition, this study will make contributions to the investigations of the influences of oxidative stress on the formation and development of CRC.
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Cromatografía Líquida de Alta Presión/métodos , Neoplasias Colorrectales/diagnóstico , Guanosina/análogos & derivados , ARN/orina , Espectrometría de Masas en Tándem/métodos , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/orina , Neoplasias Colorrectales/orina , Guanosina/química , Guanosina/orina , Humanos , Límite de Detección , Modelos Lineales , Persona de Mediana Edad , Estrés Oxidativo , ARN/química , Reproducibilidad de los ResultadosRESUMEN
The heterogeneities of colorectal cancer (CRC) lead to staging inadequately of patients' prognosis. Here, we performed a prognostic analysis based on the tumor mutational profile and explored the characteristics of the high-risk tumors. We sequenced 338 colorectal carcinomas as the training dataset, constructed a novel five-gene (SMAD4, MUC16, COL6A3, FLG and LRP1B) prognostic signature, and validated it in an independent dataset from The Cancer Genome Atlas (TCGA). Kaplan-Meier and Cox regression analyses confirmed that the five-gene signature is an independent predictor of recurrence and prognosis in patients with Stage III colon cancer. The mutant signature translated to an increased risk of death (hazard ratio = 2.45, 95% confidence interval = 1.15-5.22, p = 0.016 in our dataset; hazard ratio = 4.78, 95% confidence interval = 1.33-17.16, p = 0.008 in TCGA dataset). RNA and bacterial 16S rRNA sequencing of high-risk tumors indicated that mutations of the five-gene signature may lead to intestinal barrier integrity, translocation of gut bacteria and deregulation of immune response and extracellular related genes. The high-risk tumors overexpressed IL23A and IL1RN genes and enriched with cancer-related bacteria (Bacteroides fragilis,Peptostreptococcus, Parvimonas, Alloprevotella and Gemella) compared to the low-risk tumors. The signature identified the high-risk group characterized by gut bacterial translocation and upregulation of interleukins of the tumor microenvironment, which was worth further researching.
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Traslocación Bacteriana , Neoplasias del Colon/diagnóstico , Neoplasias del Colon/etiología , Regulación Neoplásica de la Expresión Génica , Subunidad p19 de la Interleucina-23/genética , Mutación , Microambiente Tumoral/genética , Anciano , Biomarcadores de Tumor , Neoplasias del Colon/mortalidad , Femenino , Proteínas Filagrina , Humanos , Masculino , Metagenómica , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Modelos de Riesgos Proporcionales , ARN Ribosómico 16SRESUMEN
Glioma is one of the most common malignant tumor types of the central nervous system. It is necessary to identify biomarkers and novel therapeutic targets for glioma. The purpose of the present study was to distinguish lipid biomarkers with differential expression patterns in glioma tissues and normal brain tissues by matrix assisted laser desorption/ionization (MALDI)-imaging and MALDI-time of flight (TOF)-mass spectrometry (MS). Additionally, identification of lipid biomarkers was performed to describe novel therapeutic targets for glioma treatment. A total of six tissues from three patients with glioma and three control patients with traumatic brain injury were analyzed using UltrafleXtreme MALDI-TOF/TOF. The expression levels of 15 lipid peaks were higher in the TBT samples compared with in the GBT samples. The expression levels of another 16 lipid peaks were higher in the GBT samples compared with in the TBT samples. 14 peaks were identified as sphingomyelins using MS/MS. Additional results were also obtained from experiments using the glioma cell line U373-MG. These results indicated that treatment with the drug desipramine (Desi) inhibited the accumulation of ceramide on the cell membranes of glioma U373-MG cells. Treatment with Desi inhibited the activation of insulin-like growth factor-1 receptor and inhibited the activation of proteins in the PI3K/Akt signaling pathway.
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Wilms' tumor is one of the most common malignancies in children, and early diagnosis is critical for its subsequent treatment and prognosis. Our previous study employed proteomics to investigate protein markers in the serum of Wilms' tumor children. The present study aimed to identify specific protein markers in Wilms' tumor. Proteomic comparison of Wilms' tumor with normal kidney tissues and the sera of systemic inflammatory response syndrome (SIRS) controls was performed. Surface-enhanced laser desorption ionization time-of-flight (SELDI-TOF-MS) identified a protein with m/z 8350 as specific to Wilms' tumor. The target protein was purified using sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and identified as profilin-1 by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF). Its expression was validated using real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Our data identify profilin-1 as a potential protein marker for Wilms' tumor and demonstrate the feasibility of the above procedures for screening and identification of tumor-specific protein markers.
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Proteínas de Neoplasias/metabolismo , Proteómica/métodos , Tumor de Wilms/metabolismo , Niño , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Proteínas de Neoplasias/sangre , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Tumor de Wilms/sangre , Tumor de Wilms/diagnósticoRESUMEN
We analysed the changes in viral protein expression in HBeAg-negative chronic hepatitis B (CHB). In total, 160 samples were obtained from individuals infected by hepatitis B virus (HBV) and divided into four groups. Group A included 71 cases of hepatitis B e antigen (HBeAg)-negative CHB, Group B included 58 cases of inactive seroconverters and Group C included 31 cases of HBeAg-positive CHB. Group D included 22 normal healthy individuals as a control. All serum samples were examined using surface enhance laser desorption/ionization time of flight-mass spectrometry (SELDI-TOF-MS). The results indicated that a peak with 4140 m/z increased markedly in Group A at 1295.55 ± 745.87, which was significantly different from that in Group B at 896.99 ± 534.86 (P = 0.013). This peak indicated a close relationship with HBV DNA replication and may contribute to pathogenesis of HBeAg-negative chronic hepatitis.
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Proteínas Sanguíneas/genética , Antígenos e de la Hepatitis B/sangre , Hepatitis B Crónica/sangre , Hepatitis B Crónica/genética , Adulto , Anciano , Biomarcadores , Estudios de Casos y Controles , Biología Computacional/métodos , ADN Viral , Femenino , Hepatitis B Crónica/virología , Humanos , Masculino , Persona de Mediana Edad , Reproducibilidad de los Resultados , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Carga Viral , Adulto JovenRESUMEN
In human, loss of Acid Sphingomeylinase (ASM/SMPD1) causes Niemann-Pick Disease, type A. ASM hydrolyzes sphingomyelins to produce ceramides but protein targets of ASM remain largely unclear. Our mass-spectrometry-based proteomic analyses have identified >100 proteins associated with the ASM-dependent, detergent-resistant membrane microdomains (lipid rafts), with >60% of these proteins being palmitoylated, including SNAP23, Src-family kinases Yes and Lyn, and Ras and Rab family small GTPases. Inactivation of ASM abolished the presence of these proteins in the plasma membrane, with many of them trapped in the Golgi. While palmitoylation inhibitors and palmitoylation mutants phenocopied the effects of ASM inactivation, we demonstrated that ASM is required for the transport of palmitoylated proteins, such as SNAP23 and Lyn, from the Golgi to the plasma membrane without affecting palmitoylation directly. Importantly, ASM delivered extracellularly can regulate the trafficking of SNAP23 from the Golgi to the plasma membrane. Our studies suggest that ASM, acting at the plasma membrane to produce ceramides, regulates the localization and trafficking of the palmitoylated proteins.
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[This corrects the article DOI: 10.1371/journal.pone.0167109.].
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Colorectal cancer (CRC) is a leading cause of cancer death. This study was conducted to investigate the functions and mechanisms of Zyxin (ZYX) in CRC. Multiomics analysis associated ZYX with CRC metastasis. ZYX expression levels were increased in human CRC tissues and related to shorter recurrence-free survival. Knockdown of ZYX expression resulted in inhibition of cell growth, invasion, and migration in vitro and in vivo. Comprehensive analysis of gene microarray analysis showed that ZYX may activate the pathway of NUPR1 and JNK, inhibit CST5, regulate focal adhesion (FA), and affect epithelial-mesenchymal transition in CRC cells. Results of gene microarray and membrane protein isobaric tags with relative and absolute quantitation labeling mass spectrometry found ten differentially expressed genes, which were associated with ZYX activity. Furthermore, real-time polymerase chain reaction was used to validate the expression patterns of selected genes in the integrative analysis. Taken together, our findings provide the first evidence that decreased expression level of ZYX impairs CRC cell proliferation and metastasis probably via the FA pathway.
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SOX2 is a dose-dependent master stem cell protein that controls the self-renewal and pluripotency or multipotency of embryonic stem (ES) cells and many adult stem cells. We have previously found that SOX2 protein is monomethylated at lysine residues 42 and 117 by SET7 methyltransferase to promote SOX2 proteolysis, whereas LSD1 and PHF20L1 act on both methylated Lys-42 and Lys-117 to prevent SOX2 proteolysis. However, the mechanism by which the methylated SOX2 protein is degraded remains unclear. Here, we report that L3MBTL3, a protein with the malignant-brain-tumor (MBT) methylation-binding domain, is required for SOX2 proteolysis. Our studies showed that L3MBTL3 preferentially binds to the methylated Lys-42 in SOX2, although mutation of Lys-117 also partially reduces the interaction between SOX2 and L3MBTL3. The direct binding of L3MBTL3 to the methylated SOX2 protein leads to the recruitment of the CRL4DCAF5 ubiquitin E3 ligase to target SOX2 protein for ubiquitin-dependent proteolysis. Whereas loss of either LSD1 or PHF20L1 destabilizes SOX2 protein and impairs the self-renewal and pluripotency of mouse ES cells, knockdown of L3MBTL3 or DCAF5 is sufficient to restore the protein levels of SOX2 and rescue the defects of mouse ES cells caused by LSD1 or PHF20L1 deficiency. We also found that retinoic acid-induced differentiation of mouse ES cells is accompanied by the enhanced degradation of the methylated SOX2 protein at both Lys-42 and Lys-117. Our studies provide novel insights into the mechanism by which the methylation-dependent degradation of SOX2 protein is controlled by the L3MBTL3-CRL4DCAF5 ubiquitin ligase complex.
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Proteínas de Unión al ADN/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Factores de Transcripción SOXB1/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Línea Celular , Humanos , Metilación , Ratones , Ratones Endogámicos C57BL , Células Madre Embrionarias de Ratones/citología , Células Madre Embrionarias de Ratones/metabolismo , Estabilidad Proteica , Proteolisis , UbiquitinaciónRESUMEN
Many non-histone proteins are lysine methylated and a novel function of this modification is to trigger the proteolysis of methylated proteins. Here, we report that the methylated lysine 142 of DNMT1, a major DNA methyltransferase that preserves epigenetic inheritance of DNA methylation patterns during DNA replication, is demethylated by LSD1. A novel methyl-binding protein, L3MBTL3, binds the K142-methylated DNMT1 and recruits a novel CRL4DCAF5 ubiquitin ligase to degrade DNMT1. Both LSD1 and PHF20L1 act primarily in S phase to prevent DNMT1 degradation by L3MBTL3-CRL4DCAF5. Mouse L3MBTL3/MBT-1 deletion causes accumulation of DNMT1 protein, increased genomic DNA methylation, and late embryonic lethality. DNMT1 contains a consensus methylation motif shared by many non-histone proteins including E2F1, a key transcription factor for S phase. We show that the methylation-dependent E2F1 degradation is also controlled by L3MBTL3-CRL4DCAF5. Our studies elucidate for the first time a novel mechanism by which the stability of many methylated non-histone proteins are regulated.
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ADN (Citosina-5-)-Metiltransferasa 1/genética , Proteínas de Unión al ADN/genética , Factor de Transcripción E2F1/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Complejos de Ubiquitina-Proteína Ligasa/metabolismo , Secuencias de Aminoácidos , Animales , ADN (Citosina-5-)-Metiltransferasa 1/química , ADN (Citosina-5-)-Metiltransferasa 1/metabolismo , Metilación de ADN , Proteínas de Unión al ADN/metabolismo , Factor de Transcripción E2F1/metabolismo , Femenino , Eliminación de Gen , Histona Demetilasas/genética , Histona Demetilasas/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Metilación , Ratones , Unión Proteica , Estabilidad Proteica , Proteolisis , Complejos de Ubiquitina-Proteína Ligasa/genéticaRESUMEN
The serum protein markers of colorectal adenoma in patients with a family history of colorectal cancer have been rarely reported. Serum samples from colorectal adenoma patients with or without a family history of colorectal cancer and healthy controls were profiled using Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS). The model to distinguish colorectal adenoma patients with a family history of colorectal cancer from atypical hereditary colorectal families (CRA-H) and sporadic colorectal adenoma patients without a family history of colorectal cancer (CRA-S) was established with 85.0% accuracy. The model distinguishing CRA-H from healthy individuals was established with 90.0% specificity and 86.7% sensitivity. Additionally, five peaks (2202, 5821, 3260, 2480, and 2218) showing differential expression in advanced colorectal adenoma patients with a family history of colorectal cancer were selected. The protein Kininogen 1 (KNG1) was identified in colorectal adenoma patients and validated using Western Blotting. KNG1 may be a biomarker for colorectal adenoma patients with a family history of colorectal cancer.
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Colorectal cancer (CRC) has complex pathological features that defy the linear-additive reasoning prevailing in current biomedicine studies. In pursuing a mechanistic understanding behind such complexity, we constructed a core molecular-cellular interaction network underlying CRC and investigated its nonlinear dynamical properties. The hypothesis and modelling method has been developed previously and tested in various cancer studies. The network dynamics reveal a landscape of several attractive basins corresponding to both normal intestinal phenotype and robust tumour subtypes, identified by their different molecular signatures. Comparison between the modelling results and gene expression profiles from patients collected at the second affiliated hospital of Zhejiang University is presented as validation. The numerical 'driving' experiment suggests that CRC pathogenesis may depend on pathways involved in gastrointestinal track development and molecules associated with mesenchymal lineage differentiation, such as Stat5, BMP, retinoic acid signalling pathways, Runx and Hox transcription families. We show that the multi-faceted response to immune stimulation and therapies, as well as different carcinogenesis and metastasis routes, can be straightforwardly understood and analysed under such a framework.
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Neoplasias Colorrectales/genética , Modelos Teóricos , Biología de Sistemas , Estudios de Casos y Controles , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Humanos , Dinámicas no Lineales , Procesos EstocásticosRESUMEN
Colorectal cancer (CRC) is a common malignant neoplasm worldwide. It is important to identify new biomarkers for the early detection of CRC. In this study, magnetic beads and the Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) platform were used to analyse CRC and healthy control (HC) serum samples. The CRC diagnosis pattern was established to have a specificity of 94.7% and sensitivity of 92.3% in a blind test. The candidate biomarker serine/threonine kinase 4 (STK4, also known as MST1) was identified by Tandem mass spectrometry (MS/MS) and verified with western blotting and enzyme-linked immunosorbent assay (ELISA). The results indicated that there was a higher concentration of MST1 in HC subjects than stage I CRC patients for the early detection of CRC and a lower concentration in stage IV patients than in other CRC patients. The sensitivity and specificity of MST1 combined with carcinoembryonic antigen (CEA) and faecal occult blood test (FOBT) in diagnosis of colorectal cancer were 92.3% and 100%, respectively. Additionally, low MST1 expression was associated with the poor prognosis. These results illustrate that MST1 is a potential biomarker for early detection, prognosis and prediction of distant metastasis of CRC.
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Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/metabolismo , Detección Precoz del Cáncer , Proteínas Serina-Treonina Quinasas/metabolismo , Proteómica , Biomarcadores de Tumor/metabolismo , Estudios de Casos y Controles , Neoplasias Colorrectales/patología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intracelular , Masculino , Persona de Mediana Edad , Metástasis de la Neoplasia , PronósticoRESUMEN
BACKGROUND: Chloride channel accessory 1 (CLCA1) belongs to the calcium-sensitive chloride conductance protein family, which is mainly expressed in the colon, small intestine and appendix. This study was conducted to investigate the functions and mechanisms of CLCA1 in colorectal cancer (CRC). METHODS: The CLCA1 protein expression level in CRC patients was evaluated by enzyme-linked immunosorbent assay (ELISA), immunohistochemistry (IHC), and western blotting analysis. Using CRISPR/Cas9 technology, CLCA1-upregulated (CLCA1-ACT) and CLCA1-knockout cells (CLCA1-KO), as well as their respective negative controls (CLCA1-ACT-NC and CLCA1-KO-NC), were constructed from the SW620 cell line. Cell growth and metastatic ability were assessed both in vitro and in vivo. The association of CLCA1 with epithelial-mesenchymal transition (EMT) and other signaling pathways was determined by western blotting assays. RESULTS: The expression level of CLCA1 in CRC tissues was significantly decreased compared with that in adjacent normal tissue (P< 0.05). Meanwhile, the serum concentration of CLCA1 in CRC patients was also significantly lower when compared with that of healthy controls (1.48 ± 1.06 ng/mL vs 1.06 ± 0.73 ng/mL, P = 0.0018). In addition, CLCA1 serum concentration and mRNA expression level in CRC tissues were inversely correlated with CRC metastasis and tumor stage. Upregulated CLCA1 suppressed CRC growth and metastasis in vitro and in vivo, whereas inhibition of CLCA1 led to the opposite results. Increased expression levels of CLCA1 could repress Wnt signaling and the EMT process in CRC cells. CONCLUSIONS: Our findings suggest that increased expression levels of CLCA1 can suppress CRC aggressiveness. CLCA1 functions as a tumor suppressor possibly via inhibition of the Wnt/beta-catenin signaling pathway and the EMT process.
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Canales de Cloruro/metabolismo , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Vía de Señalización Wnt , Animales , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Transformación Celular Neoplásica , Transición Epitelial-Mesenquimal , Femenino , Humanos , Masculino , RatonesRESUMEN
Breast cancer is one of the most commonly diagnosed and death-related cancers in women worldwide. Mammography is routinely used for screening and invasive examinations such as painful tissue biopsies were recommended for patients with abnormal screening outcomes. However, a considerable proportion of these cases turn out to be benign lesions. Thus, novel non-invasive approach for discriminating breast cancer from benign lesions is desirable. Herein, we applied a high-throughput ultra performance liquid chromatography-electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) analysis to determine the oxidative DNA damage biomarker, 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in urine samples from 60 patients with early-stage breast cancer (stage I, II), 51 patients with benign breast diseases and 73 healthy volunteers. We demonstrated that the concentration of urinary 8-oxodG in patients with early-stage breast cancer was significantly higher not only than that in healthy controls, but also than that in patients with benign breast diseases, whereas no significant difference of urinary 8-oxodG level was observed between benign breast diseases group and healthy control group. Moreover, there was significant difference between early-stage breast cancer group and non-cancerous group which consisted of benign breast diseases patients and healthy controls. Besides, logistic regression analysis and receiver operator characteristic (ROC) curve analysis were also performed. Our findings indicate that the marked increase of 8-oxodG in urine may serve as a potential biomarker for the risk estimation, early screening and detection of breast cancer, particularly for discriminating early-stage breast cancer from benign lesions.