An integrative approach based on GC-qMS and NMR metabolomics data as a comprehensive strategy to search potential breast cancer biomarkers.

INTRODUCTION: Globally, breast cancer (BC) is leading at the top of women's diseases and, as a multifactorial disease, there is the need for the development of new approaches to aid clinicians on monitoring BC treatments. In this sense, metabolomic studies have become an essential tool allowing the establishment of interdependency among metabolites in biological samples. OBJECTIVE: The combination of nuclear magnetic resonance (NMR) and gas chromatography-quadrupole mass spectrometry (GC-qMS) based metabolomic analyses of urine and breast tissue samples from BC patients and cancer-free individuals was used. METHODS: Multivariate statistical tools were used in order to obtain a panel of metabolites that could discriminate malignant from healthy status assisting in the diagnostic field. Urine samples (n = 30), cancer tissues (n = 30) were collected from BC patients, cancer-free tissues were resected outside the tumor margin from the same donors (n = 30) while cancer-free urine samples (n = 40) where obtained from healthy subjects and analysed by NMR and GC-qMS methodologies. RESULTS: The orthogonal partial least square discriminant analysis model showed a clear separation between BC patients and cancer-free subjects for both classes of samples. Specifically, for urine samples, the goodness of fit (R2Y) and predictive ability (Q2) was 0.946 and 0.910, respectively, whereas for tissue was 0.888 and 0.813, revealing a good predictable accuracy. The discrimination efficiency and accuracy of tissue and urine metabolites was ascertained by receiver operating characteristic curve analysis that allowed the identification of metabolites with high sensitivity and specificity. The metabolomic pathway analysis identified several dysregulated pathways in BC, including those related with lactate, valine, aspartate and glutamine metabolism. Additionally, correlations between urine and tissue metabolites were investigated and five metabolites (e.g. acetone, 3-hexanone, 4-heptanone, 2-methyl-5-(methylthio)-furan and acetate) were found to be significant using a dual platform approach. CONCLUSION: Overall, this study suggests that an improved metabolic profile combining NMR and GC-qMS may be useful to achieve more insights regarding the mechanisms underlying cancer.

Lipid biosignature of breast cancer tissues by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

PURPOSE: One of the hallmarks of cancer cells is the demand of supply for the synthesis of new membranes involved in cell proliferation and lipids have an important role in cellular structure, signaling pathways and progression of cancer. In this sense, lipid studies have become an essential tool allowing the establishment of signatures associated with breast cancer (BC). In this regard, some metabolic processes including proteins, nucleic acids and lipid synthesis are enhanced as part of cancer-associated metabolic reprogramming, as a requirement for cell growth and proliferation. METHODS: Pairwise samples of breast active carcinoma (BAC) and breast cancer-free tissues were collected from n = 28 patients and analyzed by MALDI-TOF MS. RESULTS: Major lipid species are identified in the MALDI-TOF mass spectra, with certain phosphatidylinositols (PIs) detectable only in BAC. Statistical analysis revealed significant differences (p < 0.05) between ratios lysophosphatidylcholine (LPC) 16:0/phosphatidylcholine (PC) 16:0_18:2 between AC and CF groups as well as for BC stages II and III. The ratio PC 16:0_18:2/PC16:0_18:1 was statistically different between AC and CF groups. The one-way ANOVA revealed that there are no statistical differences among BC stages (I, II and III) within AC group. Comparing BC stages, the significance impact increased (p < 0.05) with stage. CONCLUSION: The obtained data revealed MALDI-TOF MS as a powerful tool to explore lipid signatures and the enzyme activity associated with BC and possibly establish novel disease markers.

Volatomic pattern of breast cancer and cancer-free tissues as a powerful strategy to identify potential biomarkers.

Breast cancer (BC), ranked as the fifth amongst all cancers, remains at the top of women's cancers worldwide followed by colorectal, lung, cervix, and stomach cancers. The main handicap of most of the screening/diagnostic methods is based on their low sensitivity and specificity and the invasive behavior of most sampling procedures. The aim of this study was to establish the volatomic pattern of BC and cancer-free (CF) tissues (n = 30) from the same patients, as a powerful tool to identify a set of volatile organic metabolite (VOM) potential BC biomarkers which might be used together or complement with the traditional BC diagnostics strategies, through the integration of chromatographic data, obtained by solid-phase microextraction followed by gas chromatography-mass spectrometry (SPME/GC-qMS), with chemometric tools. A total of four metabolites: limonene, decanoic acid, acetic acid and furfural presented the highest contribution towards discrimination of BC and CF tissues (VIP > 1, p < 0.05). The discrimination efficiency and accuracy of BC tissue metabolites was ascertained by ROC curve analysis that allowed the identification of some metabolites with high sensitivity and specificity. The results obtained with this approach suggest the possibility of identifying endogenous metabolites as a platform to find potential BC biomarkers and pave the way to investigate the related metabolomic pathways in order to improve BC diagnostic tools. Moreover, deeper investigations could unravel novel mechanistic insights into the disease pathophysiology.

Celiac disease in children from Madeira island and its prevalence in first degree relatives.

CONTEXT: It is well recognized that celiac disease is an immune-mediated systemic disorder highly prevalent among relatives of celiac patients. OBJECTIVES: The aim of this study is to determine the prevalence of celiac disease in a group of first degree relatives of celiac children, and to access the frequency of human leukocyte antigen HLA-DQ2 and DQ8 in celiac disease patients and their affected relatives. METHODS: A survey was conducted of 39 children with celiac disease with follow-up in the Pediatric outpatient's clinic of Dr. Nélio Mendonça Hospital, in Madeira Island, Portugal. Were invited 110 first degree relatives to undergo serological screen for celiac disease with IgA antibody to human recombinant tissue transglutaminase (IgA-TGG) quantification. In all seropositive relatives, small intestinal biopsy and HLA typing was recommended. RESULTS: HLA- typing was performed in 38 celiac patients, 28/74% DQ2 positive, 1/2% DQ8 positive and 9/24% incomplete DQ2. Positive IgA-TGG was found in five out of the 95 relatives, and CD was diagnosed in three of them. Three relatives had the presence of HLA-DQ2, two were DQ2 incomplete (DQB1*02). CONCLUSIONS: The prevalence of celiac disease among first degree celiac patients´ relatives was 3.1%, 4.5 times higher than the general Portuguese population (0,7%) witch reinforces the need of extensive diagnostic screening in this specific group. HLA-DQ2 typing may be a tool in the diagnostic approach.

Celiac disease nn children from madeira island and its prevalence in first degree reaatives / Doença celíaca em idade pediátrica na Ilha da Madeira e sua prevalência em familiares de primeiro grau

Context It is well recognized that celiac disease is an immune-mediated systemic disorder highly prevalent among relatives of celiac patients. Objectives The aim of this study is to determine the prevalence of celiac disease in a group of first degree relatives of celiac children, and to access the frequency of human leukocyte antigen HLA-DQ2 and DQ8 in celiac disease patients and their affected relatives. Methods A survey was conducted of 39 children with celiac disease with follow-up in the Pediatric outpatient’s clinic of Dr. Nélio Mendonça Hospital, in Madeira Island, Portugal. Were invited 110 first degree relatives to undergo serological screen for celiac disease with IgA antibody to human recombinant tissue transglutaminase (IgA-TGG) quantification. In all seropositive relatives, small intestinal biopsy and HLA typing was recommended. Results HLA- typing was performed in 38 celiac patients, 28/74% DQ2 positive, 1/2% DQ8 positive and 9/24% incomplete DQ2. Positive IgA-TGG was found in five out of the 95 relatives, and CD was diagnosed in three of them. Three relatives had the presence of HLA-DQ2, two were DQ2 incomplete (DQB1*02). Conclusions The prevalence of celiac disease among first degree celiac patients´ relatives was 3.1%, 4.5 times higher than the general Portuguese population (0,7%) witch reinforces the need of extensive diagnostic screening in this specific group. HLA-DQ2 typing may be a tool in the diagnostic approach. .

Contexto A doença celíaca é uma doença sistémica autoimune muito prevalente nos familiares de primeiro grau de doentes celíacos. Objetivos O objetivo deste estudo é determinar a prevalência de doença celíaca, num grupo de familiares de primeiro grau de crianças com o diagnóstico de doença celíaca e, determinar a frequência de antígeno leucocitário humano (HLA)-DQ2 e DQ8 nos doentes celíacos e seus familiares afetados. Métodos Foi feita a pesquisa dos processos clínicos de 39 crianças com o diagnóstico de doença celíaca seguidas na consulta de Gastroenterologia Pediátrica do Hospital Dr. Nélio Mendonça na Ilha da Madeira, Portugal. Foram convidados 110 familiares de primeiro grau para a realização do rastreio serológico de doença celíaca através da quantificação do anticorpo IgA anti-transglutaminase tecidular humano (IgA-TGG). Aos familiares com resultado positivo no rastreio, foi recomendada a realização de biópsia intestinal e tipificação HLA. Resultados A tipificação HLA foi realizada em 38 crianças. Verificou-se a presença do heterodímero DQ2 em 28/74%, DQ8 em 1/2% e DQ2 incompleto em 9/24% das crianças. O rastreio de DC com IgA-TGG foi positivo em cinco dos 95 familiares analisados, tendo sido diagnosticada doença celíaca em três destes. Verificou-se a presença do heterodímero HLA-DQ2 em três familiares e HLA-DQ2 incompleto (DQB1*02) em dois familiares. Conclusões A prevalência de doença celíaca em familiares de primeiro grau de doentes celíacos foi 3.1%, 4.5 vezes mais elevada do que a da população Portuguesa geral (0,7%), o que reforça a importância de alargar o rastreio a este grupo específico. A tipificação ...