RESUMEN
Alkylresorcinols (ARs) are polyphenolic compounds with a wide spectrum of biological activities and are potentially involved in the regulation of host metabolism. The present study aims to establish whether ARs can be produced by the human gut microbiota and to evaluate alterations in content in stool samples as well as metabolic activity of the gut microbiota of C57BL, db/db, and LDLR (-/-) mice according to diet specifications and olivetol (5-n-pentylresorcinol) supplementation to estimate the regulatory potential of ARs. Gas chromatography with mass spectrometric detection was used to quantitatively analyse AR levels in mouse stool samples; faecal microbiota transplantation (FMT) from human donors to germ-free mice was performed to determine whether the intestinal microbiota could produce AR molecules; metagenome sequencing analysis of the mouse gut microbiota followed by reconstruction of its metabolic activity was performed to investigate olivetol's regulatory potential. A significant increase in the amounts of individual members of AR homologues in stool samples was revealed 14 days after FMT. Supplementation of 5-n-Pentylresorcinol to a regular diet influences the amounts of several ARs in the stool of C57BL/6 and LDLR (-/-) but not db/db mice, and caused a significant change in the predicted metabolic activity of the intestinal microbiota of C57BL/6 and LDLR (-/-) but not db/db mice. For the first time, we have shown that several ARs can be produced by the intestinal microbiota. Taking into account the dependence of AR levels in the gut on olivetol supplementation and microbiota metabolic activity, AR can be assumed to be potential quorum-sensing molecules, which also influence gut microbiota composition and host metabolism.
RESUMEN
Lung cancer is referred to as the second most common cancer worldwide and is mainly associated with complex diagnostics and the absence of personalized therapy. Metabolomics may provide significant insights into the improvement of lung cancer diagnostics through identification of the specific biomarkers or biomarker panels that characterize the pathological state of the patient. We performed targeted metabolomic profiling of plasma samples from individuals with non-small cell lung cancer (NSLC, n = 100) and individuals without any cancer or chronic pathologies (n = 100) to identify the relationship between plasma endogenous metabolites and NSLC by means of modern comprehensive bioinformatics tools, including univariate analysis, multivariate analysis, partial correlation network analysis and machine learning. Through the comparison of metabolomic profiles of patients with NSCLC and noncancer individuals, we identified significant alterations in the concentration levels of metabolites mainly related to tryptophan metabolism, the TCA cycle, the urea cycle and lipid metabolism. Additionally, partial correlation network analysis revealed new ratios of the metabolites that significantly distinguished the considered groups of participants. Using the identified significantly altered metabolites and their ratios, we developed a machine learning classification model with an ROC AUC value equal to 0.96. The developed machine learning lung cancer model may serve as a prototype of the approach for the in-time diagnostics of lung cancer that in the future may be introduced in routine clinical use. Overall, we have demonstrated that the combination of metabolomics and up-to-date bioinformatics can be used as a potential tool for proper diagnostics of patients with NSCLC.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/patología , Neoplasias Pulmonares/metabolismo , Metabolómica , Biomarcadores/metabolismo , Metabolismo de los LípidosRESUMEN
BACKGROUND: Clinical guidelines highly recommended the detection of potentially targetable genetic aberrations such as anaplastic lymphoma kinase (ALK) rearrangements in patients with non-small cell lung cancer (NSCLC). Few methods, such as the ALK break apart FISH assay and IHC for ALK protein, are approved for routine diagnostics. However, some challenges exist in selecting the most reliable, robust and cost-effective algorithm, especially for large-scale screening of NSCLC patients. MATERIALS AND METHODS: A total of 4002 FFPE samples from Russian patients with NSCLC were tested for ALK rearrangement using two algorithms: FISH testing only (2334 samples) and IHC screening supported by FISH in positive or equivocal cases (1546 samples). Cross validation of the methods was performed blindly in 122 specimens. All discrepant IHC/FISH cases were examined for unbalanced 5'/3'-end ALK expression and variant-specific RT PCR. RESULTS: The sensitivity and specificity of IHC compared to FISH was 100% and 99%, respectively, therefore initial IHC screening was strongly supported. The prevalence of ALK rearrangements was estimated to be 7.8% and 6.6% for the FISH and IHC/FISH groups, respectively, with significant correlations for female gender, non-smoking status and age below 60. The use of FISH after IHC screening revealed 10 additional positive cases among equivocal samples (13.4%). Seven IHC/FISH cases (0.5% of the total group) characterized as discordant were reevaluated, and four were reclassified as truly discrepant. The PCR-based investigation revealed chimeric transcripts in IHC-/FISH+ and IHC+/FISH borderline samples, while no transcript was found in two IHC+/FISH- cases. CONCLUSION: These results reveal the utility of the two-step testing algorithm for the evaluation of potentially complicated cases with preanalytic defects, providing additional information for IHC equivocal cases without a significant increase in cost. The evaluation of discrepant cases appears to be necessary to better understand ALK biology and should be included in the routine testing algorithm.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/genética , Proteínas Tirosina Quinasas Receptoras/genética , Adulto , Anciano , Anciano de 80 o más Años , Algoritmos , Quinasa de Linfoma Anaplásico , Carcinoma de Pulmón de Células no Pequeñas/enzimología , Carcinoma de Pulmón de Células no Pequeñas/patología , Femenino , Reordenamiento Génico , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ/métodos , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/patología , Masculino , Tamizaje Masivo , Persona de Mediana Edad , Prevalencia , Federación de Rusia/epidemiología , Adulto JovenRESUMEN
PURPOSE: The epidermal growth factor receptor (EGFR) is overexpressed in several tumor types, and its expression is influenced by the length of a 5'-end microsatellite repeat (CA)n: the longer the repeat, the lower the expression. Dinucleotide repeats accumulate insertion/deletion types of mutations in tumors with microsatellite instability. We designed this study to estimate the occurrence of these mutations in EGFR(CA)n and their relevance in carcinogenesis of microsatellite instability-positive colon and gastric tumors. EXPERIMENTAL DESIGN: We analyzed the frequency of EGFR(CA)n mutations in vivo in 55 colorectal and 14 gastric microsatellite instability-positive cancers, and in vitro in single-cell clone cultures of microsatellite instability-positive colon tumor cell line LS174. Single-cell clone cultures with different repeat lengths were analyzed by fluorescent-activated cell sorter for EGFR cell-surface expression. A correlation analysis was done between EGFR(CA)n mutations and mutations in KRAS, BRAF, and p53. RESULTS: Unlike single-cell clone cultures, which exhibited higher rate of deletions compared with insertions, most of EGFR(CA)n mutations in colon and gastric tumors were insertions. Longer EGFR(CA)n correlated with lower EGFR cell-surface expression in single-cell clone cultures. In colon cancers, the elongation of the repeat was associated negatively with mutations in KRAS and BRAF, but not in p53. CONCLUSIONS: The EGFR(CA)n elongation observed in tumors cannot be explained by an intrinsic property of this repeat favoring insertions versus deletions. Instead, a selection for repeat elongation occurs in microsatellite instability-positive tumors, leading to EGFR down-regulation. These findings suggest that in microsatellite instability-positive tumors current therapies targeting EGFR overexpression may have either no effect or an opposite to the expected effect.
Asunto(s)
Neoplasias del Colon/genética , Repeticiones de Dinucleótido/genética , Receptores ErbB/genética , Inestabilidad de Microsatélites , Regiones no Traducidas 5'/genética , Secuencia de Bases , Línea Celular Tumoral , Neoplasias del Colon/patología , Análisis Mutacional de ADN , Regulación hacia Abajo , Citometría de Flujo , Frecuencia de los Genes , Genes ras/genética , Genotipo , Humanos , Mutagénesis Insercional , Mutación , Poli A/genética , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , Proteínas Proto-Oncogénicas B-raf/genética , Eliminación de Secuencia , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Proteína p53 Supresora de Tumor/genéticaRESUMEN
BACKGROUND: Matrix metalloproteinases (MMPs) contribute in multiple ways to all stages of tumor development, and a number of DNA polymorphisms in the MMP genes are associated with an increased risk of cancer. We previously identified the MMP-21 gene 572C/T polymorphism leading to Ala191Val substitution within the enzyme's catalytic domain. We performed a case-control study to test association between this polymorphism and the risk of breast cancer. PATIENTS AND METHODS: 572C/T polymorphism was analyzed by RFLP method in 396 unrelated Russian females: 76 breast cancer patients and 320 disease-free blood donors. RESULTS: The frequencies of C/C, T/C and T/T genotypes in patients (69.7%, 25.0% and 5.3%) did not differ significantly from those in controls (61.9%, 34.7% and 3.4%); the polymorphism was not associated with the increased tumor size and the presence of metastases. CONCLUSION: The MMP-21 gene 572C/T polymorphism has no significant effect on the development and progression of breast cancer.