RESUMO
BACKGROUND: Breast cancer is the most common neoplasia among women in developed countries. The risk factors of breast cancer can be distinguished in modifiable and unmodifiable factors and, among the latter, genetic factors play a key role. Copy number variations (CNVs) are genetic variants that are classified as rare when present in less than 1% of the healthy population. Since rare CNVs are often cause of diseases, over the last years, their contribution in carcinogenesis has become a relevant matter of study. E2F1 is a transcriptional factor that plays an important role in regulating cell cycle and apoptosis. Its double and conflicting role is the reason why it acts both as oncogene and as tumour suppressor, depending on cell context. Since anomalies in expression or in number of copies of E2F1 have been related to several cancers, we aimed to study number of germline copies of E2F1 in women with breast cancer in order to better elucidate their contribution as predisposing factor to this tumour. METHODS: We performed, hence, a retrospective study on 222 Italian women with breast cancer recruited from October 2002 to December 2007. TaqMan CNV assay and Real-Time PCR were carried out to analyse, respectively, E2F1 CNV and E2F1 expression in the subjects of the study. Chi square test or Fisher's exact test and Student's t-test were used to calculate the frequency of CNVs and differences in continuous variables between groups, respectively. RESULTS: Intriguingly, we found that 10/222 (4.5%) women with breast cancer had more copies than controls (0/200, 0%), furthermore, the number of copies positively correlated with E2F1 gene expression in breast cancer tissue, suggesting that the constitutive gain of the gene could translate into an increased risk of genomic instability. Additionally, we found that altered E2F1 copies were present prevalently in the patients with contralateral breast cancer (20%) and all of them had a positive family history, both typically associated with hereditary cancer. CONCLUSIONS: Our findings suggest that copy number variations of E2F1 might be a susceptibility factor for breast cancer, however, further studies on large cohorts are to be performed in order to better delineate the phenotype linked to the gain of E2F1 copies.
Assuntos
Neoplasias da Mama/genética , Fator de Transcrição E2F1/genética , Idoso , Neoplasias da Mama/patologia , Variações do Número de Cópias de DNA , Feminino , Predisposição Genética para Doença , Células Germinativas , Humanos , Itália , Pessoa de Meia-Idade , Estudos RetrospectivosAssuntos
Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/genética , Predisposição Genética para Doença/genética , Células Germinativas/metabolismo , Mutação em Linhagem Germinativa , Polimorfismo de Nucleotídeo Único , Neoplasias Gástricas/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas CLOCK/genética , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Circadianas Period/genética , Adulto JovemRESUMO
AIMS: Curative surgery remains the primary form of treatment for locally advanced rectal cancer (LARC). Recent data support the use of preoperative chemoradiotherapy (pCRT) to improve the prognosis of LARC with a significant reduction of local relapse and an increase of overall survival. Unfortunately, only 20% of the patients with LARC present complete pathological response after pCRT, whereas in 20%-40%, the response is poor or absent. METHODS: We investigated the expression level of miR-194 in n=38 patients with LARC using our public microRNA (miRNA) expression dataset. miR-194 expression was further validated by real-time quantitative PCR (qRT-PCR) and in situ hybridisation (ISH). Protein-protein interaction network and pathway enrichment analysis were performed on miR-194 targets. RESULTS AND DISCUSSION: Using biopsy samples collected at diagnosis, mir-194 was significantly upregulated in patients responding to treatment (p value=0.016). The data was confirmed with qRT-PCR (p value=0.0587) and ISH (p value=0.026). Protein-protein interaction network and pathway enrichment analysis reveal a possible mechanism of susceptibility to pCRT involving Wnt pathway via its downstream mediator TRAF6. Finally, we interrogated the Comparative Toxicogenomics Database database in order to identify those chemical compounds able to mimic the biological effects of miR-194 as new possible therapeutic option in LARC treatment. The present study combining miRNA expression profiling with integrative computational biology identified miR-194 as predictive biomarker of response to pCRT. Using known and predicted drug mechanism of action, we then identified possible chemical compounds for further in vitro validation.
Assuntos
Adenocarcinoma/genética , Biomarcadores Tumorais/genética , MicroRNAs/biossíntese , Neoplasias Retais/genética , Adenocarcinoma/terapia , Adulto , Idoso , Quimiorradioterapia Adjuvante , Feminino , Humanos , Masculino , MicroRNAs/análise , Pessoa de Meia-Idade , Terapia Neoadjuvante , Neoplasias Retais/terapia , Resultado do TratamentoRESUMO
Plasmonic nanostructures show important properties for biotechnological applications, but they have to be guided on the target for exploiting their potentialities. Antibodies are the natural molecules for targeting. However, their possible adverse immunogenic activity and their cost have suggested finding other valid substitutes. Small molecules like peptides can be an alternative source of targeting agents, even if, as single molecules, their binding affinity is usually not very good. GE11 is a small dodecapeptide with specific binding to the epidermal growth factor receptor (EGFR) and low immunogenicity. The present work shows that thousands of polyethylene glycol (PEG) chains modified with lysines and functionalized with GE11 on clusters of naked gold nanoparticles, obtained by laser ablation in water, achieves a better targeting activity than that recorded with nanoparticles decorated with the specific anti-EGFR antibody Cetuximab (C225). The insertion of the cationic spacer between the polymeric part of the ligand and the targeting peptide allows for a proper presentation of GE11 on the surface of the nanosystems. Surface enhanced resonance Raman scattering signals of the plasmonic gold nanoparticles are used for quantifying the targeting activity. Molecular dynamic calculations suggest that subtle differences in the exposition of the peptide on the PEG sea are important for the targeting activity.
Assuntos
Cetuximab , Sistemas de Liberação de Medicamentos/métodos , Receptores ErbB/antagonistas & inibidores , Ouro , Nanopartículas Metálicas/química , Peptídeos , Polietilenoglicóis , Células CACO-2 , Cetuximab/química , Cetuximab/farmacologia , Receptores ErbB/metabolismo , Ouro/química , Ouro/farmacologia , Humanos , Peptídeos/química , Peptídeos/farmacologia , Polietilenoglicóis/química , Polietilenoglicóis/farmacologiaRESUMO
Intracellular calcium influences an array of pathways and affects cellular processes. With the rapidly progressing research investigating the molecular identity and the physiological roles of the mitochondrial calcium uniporter (MCU) complex, we now have the tools to understand the functions of mitochondrial Ca2+ in the regulation of pathophysiological processes. Herein, we describe the role of key MCU complex components in insulin resistance in mouse and human adipose tissue. Adipose tissue gene expression was analyzed from several models of obese and diabetic rodents and in 72 patients with obesity as well as in vitro insulin-resistant adipocytes. Genetic manipulation of MCU activity in 3T3-L1 adipocytes allowed the investigation of the role of mitochondrial calcium uptake. In insulin-resistant adipocytes, mitochondrial calcium uptake increased and several MCU components were upregulated. Similar results were observed in mouse and human visceral adipose tissue (VAT) during the progression of obesity and diabetes. Intriguingly, subcutaneous adipose tissue (SAT) was spared from overt MCU fluctuations. Furthermore, MCU expression returned to physiological levels in VAT of patients after weight loss by bariatric surgery. Genetic manipulation of mitochondrial calcium uptake in 3T3-L1 adipocytes demonstrated that changes in mitochondrial calcium concentration ([Ca2+]mt) can affect mitochondrial metabolism, including oxidative enzyme activity, mitochondrial respiration, membrane potential, and reactive oxygen species formation. Finally, our data suggest a strong relationship between [Ca2+]mt and the release of IL-6 and TNFα in adipocytes. Altered mitochondrial calcium flux in fat cells may play a role in obesity and diabetes and may be associated with the differential metabolic profiles of VAT and SAT.
Assuntos
Adipócitos/metabolismo , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Resistência à Insulina/fisiologia , Mitocôndrias/metabolismo , Células 3T3-L1 , Adulto , Animais , Estudos de Casos e Controles , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Feminino , Humanos , Gordura Intra-Abdominal/metabolismo , Gordura Intra-Abdominal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Camundongos Transgênicos , Pessoa de Meia-Idade , Mitocôndrias/patologia , Obesidade/genética , Obesidade/metabolismo , Obesidade/patologia , Estado Pré-Diabético/genética , Estado Pré-Diabético/metabolismo , Estado Pré-Diabético/patologia , Gordura Subcutânea/metabolismo , Gordura Subcutânea/patologiaRESUMO
Adenosquamous carcinoma (ASC) of the gallbladder is a rare malignant tumor that is characterized by a coexisting of glandular and squamous components. In a case of ASC, we performed hotspot multigene mutational profiling of 164 hotspot regions of eleven cancer-associated genes (AKT1, APC, BRAF, CTNNB1, KIT, KRAS, NRAS, PDGFRA, PIK3CA, PTEN and TP53) in the two microdissected components. Both tumor phenotypes resulted characterized by a p.E542K point mutation in the PIK3CA gene, whereas adenocarcinoma component revealed also a TP53 Q331* homozygous stop mutation. Of note, coexisting high-grade dysplastic epithelium was characterized by a mixed cell population, with an upper part featuring a glandular differentiation and a basal layer of p63 positive (squamous committed) cells. Overall these data provide evidence of an early squamous differentiation of the lesion with a common genetic landscape of the two components.
Assuntos
Carcinoma Adenoescamoso/genética , Neoplasias da Vesícula Biliar/genética , Idoso , Feminino , Perfilação da Expressão Gênica , Humanos , TranscriptomaRESUMO
BACKGROUND: The number of studies on the association between clock genes' polymorphisms and cancer susceptibility has increased over the last years but the results are often conflicting and no comprehensive overview and quantitative summary of the evidence in this field is available. RESULTS: Literature search identified 27 eligible studies comprising 96756 subjects (cases: 38231) and investigating 687 polymorphisms involving 14 clock genes. Overall, 1025 primary and subgroup meta-analyses on 366 gene variants were performed. Study distribution by tumor was as follows: breast cancer (n=15), prostate cancer (n=3), pancreatic cancer (n=2), non-Hodgkin's lymphoma (n=2), glioma (n=1), chronic lymphocytic leukemia (n=1), colorectal cancer (n=1), non-small cell lung cancer (n=1) and ovarian cancer (n=1).We identified 10 single nucleotide polymorphisms (SNPs) significantly associated with cancer risk: NPAS2 rs10165970 (mixed and breast cancer shiftworkers), rs895520 (mixed), rs17024869 (breast) and rs7581886 (breast); CLOCK rs3749474 (breast) and rs11943456 (breast); RORA rs7164773 (breast and breast cancer postmenopausal), rs10519097 (breast); RORB rs7867494 (breast cancer postmenopausal), PER3 rs1012477 (breast cancer subgroups) and assessed the level of quality evidence to be intermediate. We also identified polymorphisms with lower quality statistically significant associations (n=30). CONCLUSIONS: Our work supports the hypothesis that genetic variation of clock genes might affect cancer risk. These findings also highlight the need for more efforts in this research field in order to fully establish the contribution of clock gene variants to the risk of developing cancer. METHODS: We conducted a systematic review and meta-analysis of the evidence on the association between clock genes' germline variants and the risk of developing cancer. To assess result credibility, summary evidence was graded according to the Venice criteria and false positive report probability (FPRP) was calculated to further validate result noteworthiness. Subgroup meta-analysis was also performed based on participant features and tumor type. The breast cancer subgroup was further stratified by work conditions, estrogen receptor/progesterone receptor status and menopausal status, conditions associated with the risk of breast cancer in different studies.
Assuntos
Relógios Circadianos/genética , Adulto , Feminino , Variação Genética , Humanos , Neoplasias/patologia , Polimorfismo de Nucleotídeo Único , Fatores de RiscoRESUMO
Colorectal cancer is characterized by aberrant Cyclooxigenase-2 (COX-2) and ß-Catenin pathways. Recently, the protease inhibitor SerpinB3 has been described overexpressed in more advanced stages of this tumor. Aim of the study was to explore the possible relationship between these molecules in this setting. We evaluated colorectal cancer specimens from 105 patients and a positive correlation between SerpinB3, COX-2 and ß-Catenin expression was observed, with higher levels in tumor than in adjacent tissue. The highest levels were associated with pathologic parameters of poor prognosis, including vascular invasion, lymph node metastasis and perineural invasion. The molecular and protein profiles of COX-2 and ß-Catenin were analyzed in cell lines with different expression of SerpinB3. In those with high expression of SerpinB3, COX-2 and ß-Catenin were higher than in controls. Cells with high levels of SerpinB3 showed higher proliferation and invasion compared to controls. In conclusion, in colorectal cancer SerpinB3, COX-2 and ß-Catenin are positively correlated and associated with more advanced tumor stage. The in vitro experimental results support a driving role of SerpinB3 in the upregulation of COX-2/ ß-Catenin positive loop, associated with a more aggressive cellular phenotype.
Assuntos
Antígenos de Neoplasias/genética , Neoplasias Colorretais/genética , Ciclo-Oxigenase 2/genética , Regulação Neoplásica da Expressão Gênica , Serpinas/genética , Regulação para Cima , beta Catenina/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Ciclo-Oxigenase 2/metabolismo , Feminino , Células HT29 , Células Hep G2 , Humanos , Immunoblotting , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Masculino , Microscopia de Fluorescência , Pessoa de Meia-Idade , Prognóstico , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Serpinas/metabolismo , Transdução de Sinais/genética , beta Catenina/metabolismoRESUMO
BACKGROUND: Insulin-resistance and hyperinsulinemia could have a role in the growing incidence of esophageal adenocarcinoma (EAC) and its pre-cancerous lesion Barrett's Esophagus (BE). HER2 activation has also a pivotal role in EAC carcinogenesis but no data correlate these two phenomena in this disease context. AIMS: To investigate the role of hyperinsulinemia in BE-dysplasia-adenocarcinoma sequence and the possible relationship between insulin-mediated and HER2 signaling in EAC development. METHODS: Serum insulin, C-peptide, IGF1, glucagon, IL-6, TNF-alpha, leptin, adiponectin and Insulin-Resistance-index were analyzed in 19 patients with gastro-esophageal reflux disease, 51 with BE, 24 with dysplastic-BE and 14 with EAC. Insulin/IGF1/HER2 pathways were analyzed in esophageal biopsies using Luminex® Technology. Insulin effect was also evaluated in EAC-derived OE19 cells. Data were analyzed by Fisher's exact test, Kruskal-Wallis test, Mann-Whitney U-test, Cuzick's test and Spearman correlation coefficient calculation. RESULTS: Insulin-Resistance-index, insulin and C-peptide levels increased along with disease progression (p=0.019, p=0.002, p<0.0001, respectively) and correlated with HER2 expression and with downstream mediators phospho-Akt and phospho-mTOR in esophageal tissue. In vitro, insulin was also able to induce cell proliferation through HER2 activation. CONCLUSIONS: Our data pinpoint a possible role of hyperinsulinemia in the Barrett's Esophagus metaplasia-dysplasia-adenocarcinoma sequence through HER2 activation in esophageal epithelial cells.
Assuntos
Adenocarcinoma/patologia , Esôfago de Barrett/patologia , Neoplasias Esofágicas/patologia , Insulina/sangue , Receptor ErbB-2/metabolismo , Transdução de Sinais , Adulto , Idoso , Peptídeo C/sangue , Progressão da Doença , Esôfago/patologia , Feminino , Refluxo Gastroesofágico/patologia , Humanos , Imuno-Histoquímica , Resistência à Insulina , Interleucina-6/metabolismo , Itália , Masculino , Pessoa de Meia-Idade , Serina-Treonina Quinases TOR/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Circulating cell-free DNA (cfDNA) has been considered an interesting diagnostic/prognostic plasma biomarker in tumor-bearing subjects. In cancer patients, cfDNA can hypothetically derive from tumor necrosis/apoptosis, lysed circulating cells, and some yet unrevealed mechanisms of active release. This study aimed to preliminarily analyze cfDNA in dogs with canine mammary tumors (CMTs). Forty-four neoplastic, 17 non-neoplastic disease-bearing, and 15 healthy dogs were recruited. Necrosis and apoptosis were also assessed as potential source of cfDNA on 78 CMTs diagnosed from the 44 dogs. The cfDNA fragments and integrity index significantly differentiated neoplastic versus non-neoplastic dogs (P<0.05), and allowed the distinction between benign and malignant lesions (P<0.05). Even if without statistical significance, the amount of cfDNA was also affected by tumor necrosis and correlated with tumor size and apoptotic markers expression. A significant (P<0.01) increase of Bcl-2 in malignant tumors was observed, and in metastatic CMTs the evasion of apoptosis was also suggested. This study, therefore, provides evidence that cfDNA could be a diagnostic marker in dogs carrying mammary nodules suggesting that its potential application in early diagnostic procedures should be further investigated.
Assuntos
DNA de Neoplasias/sangue , Doenças do Cão/sangue , Neoplasias Mamárias Animais/sangue , Animais , DNA de Neoplasias/genética , Doenças do Cão/diagnóstico , Doenças do Cão/genética , Cães , Feminino , Neoplasias Mamárias Animais/diagnóstico , Neoplasias Mamárias Animais/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismoRESUMO
In the history of medicine, nature has represented the main source of medical products. Indeed, the therapeutic use of plants certainly goes back to the Sumerian and Hippocrates and nowadays nature still represents the major source for new drugs discovery. Moreover, in the cancer treatment, drugs are either natural compounds or have been developed from naturally occurring parent compounds firstly isolated from plants and microbes from terrestrial and marine environment. A critical element of an anticancer drug is represented by its severe toxicities and, after administration, the drug concentrations have to remain in an appropriate range to be effective. Anyway, the drug dosage defined during the clinical studies could be inappropriate for an individual patient due to differences in drug absorption, metabolism and excretion. For this reason, personalized medicine, based on therapeutic drug monitoring (TDM), represents one of most important challenges in cancer therapy. Mass spectrometry sensitivity, specificity and fastness lead to elect this technique as the Golden Standard for pharmacokinetics and drug metabolism studies therefore for TDM. This review focuses on the mass spectrometry-based methods developed for pharmacokinetic quantification in human plasma of anticancer drugs derived from natural sources and already used in clinical practice. Particular emphasis was placed both on the pre-analytical and analytical steps, such as: sample preparation procedures, sample size required by the analysis and the limit of quantification of drugs and metabolites to give some insights on the clinical practice applicability. © 2015 Wiley Periodicals, Inc. Mass Spec Rev. 36:213-251, 2017.
Assuntos
Antineoplásicos/farmacocinética , Produtos Biológicos/farmacocinética , Monitoramento de Medicamentos/métodos , Espectrometria de Massas/métodos , Animais , Antineoplásicos/sangue , Produtos Biológicos/sangue , Cromatografia Líquida/instrumentação , Cromatografia Líquida/métodos , Monitoramento de Medicamentos/instrumentação , Desenho de Equipamento , Humanos , Espectrometria de Massas/instrumentação , Neoplasias/tratamento farmacológicoRESUMO
Preoperative chemoradiotherapy (pCRT) followed by surgery is the standard treatment for locally advanced rectal cancer (LARC). However, tumor response to pCRT is not uniform, and there are no effective predictive methods. This study investigated whether specific gene and miRNA expression are associated with tumor response to pCRT. Tissue biopsies were obtained from patients before pCRT and resection. Gene and miRNA expression were analyzed using a one-color microarray technique that compares signatures between responders (R) and non-responders (NR), as measured based on tumor regression grade. Two groups composed of 38 "exploration cohort" and 21 "validation cohort" LARC patients were considered for a total of 32 NR and 27 R patients. In the first cohort, using SAM Two Class analysis, 256 genes and 29 miRNAs that were differentially expressed between the NR and R patients were identified. The anti-correlation analysis showed that the same 8 miRNA interacted with different networks of transcripts. The miR-630 appeared only with the NR patients and was anti-correlated with a single transcript: RAB5B. After PAM, the following eight transcripts were strong predictors of tumor response: TMEM188, ITGA2, NRG, TRAM1, BCL2L13, MYO1B, KLF7, and GTSE1. Using this gene set, an unsupervised cluster analysis was applied to the validation cohort and correctly assigned the patients to the NR or R group with 85.7% accuracy, 90% sensitivity, and 82% specificity. All three parameters reached 100% when both cohorts were considered together. In conclusion, gene and miRNA expression profiles may be helpful for predicting response to pCRT in LARC patients. J. Cell. Physiol. 232: 426-435, 2017. © 2016 Wiley Periodicals, Inc.
Assuntos
Adenocarcinoma/genética , Adenocarcinoma/terapia , Quimiorradioterapia , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Cuidados Pré-Operatórios , Neoplasias Retais/genética , Neoplasias Retais/terapia , Adulto , Idoso , Análise por Conglomerados , Estudos de Coortes , Feminino , Perfilação da Expressão Gênica , Ontologia Genética , Humanos , Masculino , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Estadiamento de Neoplasias , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reprodutibilidade dos Testes , Resultado do TratamentoRESUMO
Colorectal cancer (CRC) whit more than a million of new cases per year is one of the most common registered cancers worldwide with few treatment options especially for advanced and metastatic patients.The tumor microenvironment is composed by extracellular matrix (ECM), cells, and interstitial fluids. Among all these constituents, in the last years an increased interest around the ECM and its potential role in cancer tumorigenesis is arisen. During cancer progression the ECM structure and composition became disorganized, allowing cellular transformation and metastasis. Up to now, the focus has mainly been on the characterization of CRC microenvironment analyzing separately structural ECM components or cell secretome modifications. A more extensive view that interconnects these aspects should be addressed. In this review, biochemical (secretome) and biomechanical (structure and architecture) changes of tumor microenvironment will be discussed, giving suggestions on how these changes can affect cancer cell behavior. J. Cell. Physiol. 232: 967-975, 2017. © 2016 Wiley Periodicals, Inc.
Assuntos
Neoplasias Colorretais/patologia , Matriz Extracelular/metabolismo , Microambiente Tumoral , Colo/patologia , HumanosRESUMO
Circulating cell-free DNA (cfDNA) was found in increased amounts in cancer patients and tumor-associated molecular alteration can be detected in cancer patient's samples. For this reason, the cfDNA analysis is actually considered as a new concept of liquid biopsy. We evaluated the presence and integrity of plasma cfDNA by ALU-based qPCR and the methylation profile of OSMR and SFRP1 genes promoter in a large cohort of colorectal cancer (CRC) patients (n = 114) in comparison to healthy subjects (n = 56) and patients with adenomatous lesions (n = 22). Moreover, we studied the prognosis value focusing on histopathological staging and survival. The cfDNA concentration and the integrity index were increased in CRC patients. The ALU83 and ALU244 fragment dosage showed a moderate discriminant capacity between CRC patients and controls and CRC and adenoma patients. Especially, cfDNA was significantly higher in CRC patients at advanced histopathological stage. In addition, the increased cfDNA level was associated with poor prognosis. A comparison of methylation profile in matched tissue and plasma on 25 CRC patients was performed and only three mismatched cases were observed. A lower methylation quantification was observed in cfDNA than tissue DNA. The cfDNA methylation frequency was statistically different in controls, adenoma and CRC patients and this frequency increased with the histopathological stage of tumor. The adenoma and CRC patients methylated cfDNA showed a higher quantity of ALU83 and ALU244. An integrated approach, combining the detection of ALU fragments and cancer type-specific epigenetic alteration, can improve diagnostic efficiency and better define the prognostic value for CRC disease.
Assuntos
Elementos Alu/genética , Biomarcadores Tumorais/sangue , Neoplasias Colorretais/sangue , DNA de Neoplasias/sangue , Idoso , Neoplasias Colorretais/patologia , Metilação de DNA/genética , DNA de Neoplasias/genética , Epigênese Genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Regiões Promotoras GenéticasRESUMO
Colorectal cancer (CRC) is one of the most common tumors in developed countries. The five-year survival rate decreases depending on how advanced the CRC is when first diagnosed. Screening has been proven to greatly reduce mortality from colorectal cancer, but an ideal screening tool is far from being established. Here, we aimed to discover and validate early CRC biomarkers by means of an untargeted/targeted metabolomic approach. A preliminary untargeted analysis of plasma lipids performed on a small patient cohort (30 plasma samples) revealed some alterations that occurred in the presence of this tumor. In particular, medium-chain fatty acids with between six and twelve carbon atoms (C6-C12) were found to be the lipid class that showed the most marked changes upon the development of CRC. In order to evaluate the utility of this lipid class as diagnostic CRC biomarkers, a further study based on a wider cohort of patients (117 plasma samples) was performed. Using a targeted approach, these fatty acids were quantified in plasma samples by means of fast gas chromatography coupled to a time-of-flight analyzer. Plasma samples from patients with CRCs at different tumor stages were analyzed and compared to those from healthy subjects, ulcerative colitis patients, high-grade dysplasia adenoma patients, and breast cancer patients in order to test the specificity and sensitivity of these possible biomarkers. Results revealed significant differences among the considered groups in terms of their C6, C8, C10, and C12 fatty acid plasma concentrations. In particular, receiver operating characteristic (ROC) curves obtained for the C10 fatty acid gave an area under the curve of 0.8195 along with a sensitivity of 87.8 % and a specificity of 80 %, strongly suggesting that it could be a valuable early diagnostic biomarker of CRC.
Assuntos
Neoplasias Colorretais/sangue , Ácidos Decanoicos/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/sangue , Estudos de Coortes , Neoplasias Colorretais/diagnóstico , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Limite de Detecção , Masculino , Pessoa de Meia-IdadeRESUMO
Irinotecan is a widely used antineoplastic drug, mostly employed for the treatment of colorectal cancer. This drug is a feasible candidate for therapeutic drug monitoring due to the presence of a wide inter-individual variability in the pharmacokinetic and pharmacodynamic parameters. In order to determine the drug concentration during the administration protocol, we developed a quantitative MALDI-MS method using CHCA as MALDI matrix. Here, we demonstrate that MALDI-TOF can be applied in a routine setting for therapeutic drug monitoring in humans offering quick and accurate results. To reach this aim, we cross validated, according to FDA and EMA guidelines, the MALDI-TOF method in comparison with a standard LC-MS/MS method, applying it for the quantification of 108 patients' plasma samples from a clinical trial. Standard curves for irinotecan were linear (R (2) ≥ 0.9842) over the concentration ranges between 300 and 10,000 ng/mL and showed good back-calculated accuracy and precision. Intra- and inter-day precision and accuracy, determined on three quality control levels were always <12.8 % and between 90.1 and 106.9 %, respectively. The cross-validation procedure showed a good reproducibility between the two methods, the percentage differences within 20 % in more than 70 % of the total amount of clinical samples analysed.
Assuntos
Camptotecina/análogos & derivados , Neoplasias Colorretais/sangue , Monitoramento de Medicamentos/métodos , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Algoritmos , Antineoplásicos/administração & dosagem , Antineoplásicos/sangue , Antineoplásicos/farmacocinética , Camptotecina/administração & dosagem , Camptotecina/sangue , Camptotecina/farmacocinética , Neoplasias Colorretais/tratamento farmacológico , Humanos , Irinotecano , Taxa de Depuração Metabólica , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Therapeutic management of Locally Advanced Rectal Cancer (LARC) involves pre-operative chemoradiotherapy (pCRT) followed by surgery. However, after pCRT the complete pathological response is approximately 20%, whereas in 20 to 40% of patients the response is poor or absent. METHODS: Cancer biopsy specimens (n= 38) and serum samples (n= 34) obtained before pCRT from 38 LARC patients were included in the study. Patients were classified in responders (R, tumor regression grade [TRG] 1-2; n= 16) and non-responders (NR, TRG 3-5; n= 22) according to the pathological response observed upon surgery. We performed miRNA microarrays analysis on biopsy specimens, and validated the selected candidates both by qRT-PCR (tissue and serum) and by in situ hybridization (tissue, miR-125b) analyses. RESULTS: Eleven miRNAs were significantly different between R and NR (miR-154, miR-409-3p, miR-127-3p, miR-214*, miR-299-5p and miR-125b overexpressed in NR; miR-33a, miR-30e, miR-338-3p, miR-200a and miR-378 decreased). In particular, miR-125b resulted to be the best candidate to discriminate the two groups (AUC of 0.9026; 95% CI, 0.7618-1.043). Additionally, miR-125b serum levels were significantly overexpressed in NR patients compared to R (p-value=0.0087), with an excellent discriminating power (AUC of 0.782; 95% CI, 0.6123-0.9518). CONCLUSIONS: The obtained results further support the clinical impact of miRNA analysis. High miR-125b expression in tissue and serum were associated with a poor treatment response in LARC patients, therefore miR-125b could be considered as a possible novel non-invasive biomarker of response in LARC treatment.
Assuntos
Adenocarcinoma/genética , Biomarcadores Tumorais/genética , MicroRNAs/genética , Neoplasias Retais/genética , Adenocarcinoma/sangue , Adenocarcinoma/terapia , Adulto , Idoso , Biomarcadores Tumorais/sangue , Quimiorradioterapia , Análise por Conglomerados , Feminino , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Humanos , Hibridização In Situ , Masculino , MicroRNAs/sangue , Pessoa de Meia-Idade , Período Pré-Operatório , Neoplasias Retais/sangue , Neoplasias Retais/terapia , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Resultado do TratamentoRESUMO
BACKGROUND: Familial adenomatous polyposis (FAP) is one of the most important clinical forms of inherited susceptibility to colorectal cancer. So far, no accepted prognostic markers are present to monitor patients with FAP. Consequently, the major problem in managing patients with FAP is the difficulty to predict when the switch between adenoma and malignant carcinoma occurs, leading to the necessity of preventive surgery. Proteomics is one of the most suitable approaches to identify biomarkers, and it is widely used in cancer research. In this investigation, we studied the circulating plasma peptides in samples collected from patients with FAP and compared the obtained results with adenoma, colorectal cancer, and control samples to discover peptides able to distinguish different phenotypes. MATERIALS AND METHODS: The peptide fingerprint was obtained by matrix-assisted laser desorption/ionization coupled to time-of-flight mass spectrometry. After statistical analysis, a subset of 45 ionic species was found differently expressed in the 4 groups considered, 12 of them peculiar to patients with FAP. Moreover, 4 ionic species were found significantly changed in the switch between adenoma and malignant carcinoma. RESULTS: Potentially prognostic peptides identified by this study derive mainly from circulating proteins, some of which are involved in the inflammatory response, such as complement C3 and C4 subjected to an exoprotease activity that seemed pathology related. CONCLUSIONS: In this study, we defined for the first time a specific panel of peptides for monitoring patients with FAP that could be profitably used to monitor and predict the pathologic evolution in adenocarcinoma malignancy.
Assuntos
Polipose Adenomatosa do Colo/sangue , Biomarcadores Tumorais/sangue , Detecção Precoce de Câncer/métodos , Mapeamento de Peptídeos/métodos , Peptídeos/sangue , Polipose Adenomatosa do Colo/diagnóstico , Adulto , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteômica/métodos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
The use of sentinel node biopsy (SNB) after neoadjuvant chemotherapy (NAC) in patients with locally advanced breast cancer is debated. Our aim was to quantitatively review the available evidence on the performance of SNB after NAC in patients with locally advanced breast cancer. We performed a systematic review (by searching the PubMed, Cochrane and Scopus databases) and random effects meta-analysis to investigate on the feasibility and accuracy of SNB in these patients. The two outcomes of interest were the sentinel node identification rate (SIR) and the false negative rate (FNR). Sensitivity analysis and meta-regression were used to investigate the potential sources of between-study heterogeneity. We retrieved 72 eligible studies enrolling 7,451 patients. Upon meta-analysis, summary SIR resulted 89.6% [95% confidence interval (CI): 87.8-91.2; heterogeneity I(2): 76.9%], which poorly compares with the 95% SIR observed in some recent series of early breast cancer. The summary FNR resulted 14.2% (CI: 12.5-16.0; heterogeneity I(2): 29.1%), which was significantly higher than the 8-10% reference value. Considering an average post-NAC lymph node positivity rate of 50%, the downstaging due to false negative SNB would occur in 7/100 patients (with an excess error rate of 2-3/100 as compared to the early-stage setting). No plausible source of between-study heterogeneity was found. Based on the largest series of studies ever meta-analyzed, our findings highlight the limits of SNB performance in this population, where the impact of SNB on patient survival is still to be defined.
Assuntos
Neoplasias da Mama/patologia , Metástase Linfática/diagnóstico , Biópsia de Linfonodo Sentinela , Neoplasias da Mama/tratamento farmacológico , Quimioterapia Adjuvante , Feminino , Humanos , Terapia NeoadjuvanteRESUMO
Early detection of colorectal cancer (CRC) remains a challenge. It has been highlighted that the pathological alterations within an organ and tissues might be reflected in serum or plasma proteomic/peptidic patterns. The aim of the study was to follow the changes in the plasma peptides associated to colorectal cancer progression by mass spectrometry. This study included 27 adenoma, 67 CRC (n = 33 I-II stage and n = 34 III-IV stage), 23 liver metastasis from CRC patients and 34 subjects disease-free as controls. For plasma peptides analysis, samples purification was performed on the Nanoporous Silica Chips technology followed by matrix-assisted laser desorption/ionisation-time of flight analysis. Since the high complexity of the obtained dataset, multivariate statistical analysis, and discriminant pattern recognition were performed for study groups classification. Forty-four of 88 ionic species were successfully identified as fragments of peptides and proteins physiologically circulating in the blood and belonging to immune and coagulation systems and inflammatory mediators. Many peptides clustered into sets of overlapping sequences with ladder-like truncation clearly associated to proteolytic processes of both endo- and exoproteases activity. Comparing to controls, a different median ion intensity of the group-type fragments distribution was observed. Moreover, the degradation pattern obtained by proteolytic cleavage was different into study groups. This pattern was specific and characteristic of each group: controls, colon tumour disease (including adenoma and CRC), and liver metastasis, revealing a role as biomarker in early diagnosis and prognosis. Our findings highlighted peculiar changes in protease activity characteristic of CRC progression from pre-cancer lesion to metastatic disease. J. Cell. Physiol. 231: 915-925, 2016. © 2015 Wiley Periodicals, Inc.