Metabolic Signatures of Lung Cancer in Biofluids: NMR-Based Metabonomics of Blood Plasma

In this work, the variations in the metabolic profile of blood plasma from lung cancer patients and healthy controls were investigated through NMR-based metabonomics, to assess the potential of this approach for lung cancer screening and diagnosis. PLS-DA modeling of CPMG spectra from plasma, subjected to Monte Carlo Cross Validation, allowed cancer patients to be discriminated from controls with sensitivity and specificity levels of about 90%. Relatively lower HDL and higher VLDL + LDL in the patients' plasma, together with increased lactate and pyruvate and decreased levels of glucose, citrate, formate, acetate, several amino acids (alanine, glutamine, histidine, tyrosine, valine), and methanol, could be detected. These changes were found to be present at initial disease stages and could be related to known cancer biochemical hallmarks, such as enhanced glycolysis, glutaminolysis, and gluconeogenesis, together with suppressed Krebs cycle and reduced lipid catabolism, thus supporting the hypothesis of a systemic metabolic signature for lung cancer. Despite the possible confounding influence of age, smoking habits, and other uncontrolled factors, these results indicate that NMR-based metabonomics of blood plasma can be useful as a screening tool to identify suspicious cases for subsequent, more specific radiological tests, thus contributing to improved disease management.ERDF - Competitive Factors Thematic Operational ProgrammeFCT/PTDC/ QUI/68017/2006FCOMP-01-0124-FEDER-007439SFRH/BD/ 63430/2009National UNESCO Committee - L'Oréal Medals of Honor for Women in Science 200Portuguese National NMR Network - RNRM

Use of Fourier-transform infrared spectroscopy to determine immunoglobulin status in camelid and equine species

Measurement of systemic immunoglobulin (Ig) concentrations in horses and camelids is important for early and accurate diagnosis of immunodeficiencies in order to provide proper medical intervention. As a consequence, there is a demand for up-to-date, economic, rapid and precise diagnostic assays for Igs. Accordingly, multiple methods for the evaluation of Ig concentrations have been evaluated in equine and camelid species, each with particular advantages and disadvantages. Fourier-transform infrared (FTIR) spectroscopy has recently emerged as a powerful diagnostic tool for the quantitative characterization of biological fluids in human and veterinary medicine. In particular, FTIR spectroscopy has proven to be an accurate, economical and reagent-free method for immunoglobulin G (IgG) quantitation in horses. Further research to investigate its potential application in the quantitation of other equine Ig isotypes and IgG subclasses is warranted. Additionally, as rapid quantitative assays for the measurement of camelid serum IgG are limited, further work to assess the use of FTIR spectroscopy in this area is desirable. The objectives of this thesis were: (1) to develop an FTIR-based assay for the measurement of IgG concentrations in alpaca serum and to compare its performance to that of the radial immunodiffusion (RID) assay, and (2) to develop FTIR-based assays for the measurement IgGa, IgGb, IgG(T), IgA, and IgM for equine plasma using ELISA assays as reference tests. The first objective of this thesis was achieved by performing RID IgG assays and collecting FTIR spectra for 175 alpaca serum samples. A FTIR-based assay was built using partial least squares regression to convert the spectroscopic data into quantitative IgG values which were compared to the RID results. Correlation coefficients and scatter plots indicated good to excellent levels of agreement between the assays. The results suggest that FTIR spectroscopy may be a useful method for IgG measurement in alpaca serum. For the second objective, IgGa, IgGb, IgG(T), IgA, and IgM concentrations were determined by ELISA assays and FTIR spectra were collected for 100 equine plasma samples. The spectra were randomly divided into training and prediction sets. The training set was used to build a calibration model for each Ig isotype or IgG subclass using partial least squares regression, while the prediction set was used to test the performance of the developed models. Pearson correlation coefficients and scatter plots displayed moderate to good agreement between FTIR and ELISA IgGb assay results but poor overall agreement for IgGa, IgG(T), IgA and IgM assay results. As well, significant differences were noted between the ELISA results of this study and the reviewed studies from the published literature. At present, a FTIR spectroscopic approach is an inaccurate technique for the measurement of Ig isotypes or IgG subclasses in equine plasma

Discrimination of healthy and colorectal cancer patients using FTIR and PLS-DA

Spectroscopic methods have already been used as effective tools in several studies involving the detection of cancer. Fourier transform infrared spectroscopy (FTIR) has already been applied in the discrimination of cancer cells and tissues or blood of patients with the disease, observing that this technique requires the use of chemometric algorithms to obtain such results. The aim of this study was to employ a partial least squares discriminant analysis (PLS-DA) with FTIR data in the discrimination of plasma samples from patients with colorectal cancer (RCC) and healthy individuals. Multivariate analysis was performed using PLS-DA of the sample triplicates (n=90) with different types of processing. The best PLS-DA condition was obtained using the 1st derivative, 1 orthogonal signal correction (OSC) and no pre-processing. With 1 factor only, the model presented a mean square error of cross-validation (RMSECV) of 0.0004 and coefficient of determination (r^2) of 1.0000. The accuracy, precision and sensitivity of the model were 100%

PESI-MS for Diagnostic Cytology

Objectives: Cytology and histology are 2 indispensable diagnostic tools for cancer diagnosis, which are rapidly increasing in importance with aging populations. We applied mass spectrometry (MS) as a rapid approach for swiftly acquiring nonmorphological information of interested cells. Conventional MS, which primarily rely on promoting ionization by pre-applying a matrix to cells, has the drawback of time-consuming both on data acquisition and analysis. As an emerging method, probe electrospray ionization-MS (PESI-MS) with a dedicated probe is capable to pierce sample and measure specimen in small amounts, either liquid or solid, without the requirement for sample pretreatment. Furthermore, PESI-MS is timesaving compared to the conventional MS. Herein, we investigated the capability of PESI-MS to characterize the cell types derived from the respiratory tract of human tissues. Study Design: PESI-MS analyses with DPiMS-2020 were performed on various type of cultured cells including 5 lung squamous cell carcinomas, 5 lung adenocarcinomas, 5 small-cell carcinomas, 4 malignant mesotheliomas, and 2 normal controls. Results: Several characteristic peaks were detected at around m/z 200 and 800 that were common in all samples. As expected, partial least squares-discriminant analysis of PESI-MS data distinguished the cancer cell types from normal control cells. Moreover, distinct clusters divided squamous cell carcinoma from adenocarcinoma. Conclusion: PESI-MS presented a promising potential as a novel diagnostic modality for swiftly acquiring specific cytological information

Multivariate classification of gene expression microarray data

L'expressiódels gens obtinguts de l'anàliside microarrays s'utilitza en molts casos, per classificar les cèllules. En aquestatesi, unaversióprobabilística del mètodeDiscriminant Partial Least Squares (p-DPLS)s'utilitza per classificar les mostres de les expressions delsseus gens. p-DPLS esbasa en la regla de Bayes de la probabilitat a posteriori. Aquestsclassificadorssónforaçats a classficarsempre.Per superaraquestalimitaciós'haimplementatl'opció de rebuig.Aquestaopciópermetrebutjarlesmostresamb alt riscd'errors de classificació (és a dir, mostresambigüesi outliers).Aquestaopció de rebuigcombinacriterisbasats en els residuals x, el leverage ielsvalorspredits. A més,esdesenvolupa un mètode de selecció de variables per triarels gens mésrellevants, jaque la majoriadels gens analitzatsamb un microarraysónirrellevants per al propòsit particular de classificacióI podenconfondre el classificador. Finalment, el DPLSs'estenen a la classificació multi-classemitjançant la combinació de PLS ambl'anàlisidiscriminant lineal

Near-infrared spectroscopy as a tool for in vivo analysis of human muscles

Recent advances in materials and fabrication techniques provided portable, performant, sensing optical spectrometers readily operated by user-friendly cabled or wireless systems. Such systems allow rapid, non-invasive, and not destructive quantitative analysis of human tissues. This proof-of-principle investigation tested whether infrared spectroscopy techniques, currently utilized in a variety of areas, could be applied in living humans to categorize muscles. Using an ASD FieldSpec\uae 4 Standard-Res Spectroradiometer with a spectral sampling capability of 1.4 nm at 350\u20131000 nm and 1.1 nm at 1001\u20132500 nm, we acquired reflectance spectra in visible short-wave infra-red regions (350\u20132500 nm) from the upper limb muscles (flexors and extensors) of 20 healthy subjects (age 25\u201389 years, 9 women). Spectra off-line analysis included preliminary preprocessing, Principal Component Analysis, and Partial Least-Squares Discriminant Analysis. Near-infrared (NIR) spectroscopy proved valuable for noninvasive assessment of tissue optical properties in vivo. In addition to the non-invasive detection of tissue oxygenation, NIR spectroscopy provided the spectral signatures (ie, \u201cfingerprints\u201d) of upper limb flexors and extensors, which represent specific, accurate, and reproducible measures of the overall biological status of these muscles. Thus, non-invasive NIR spectroscopy enables more thorough evaluation of the muscular system and optimal monitoring of the effectiveness of therapeutic or rehabilitative interventions