Information retrieval using machine learning for database curation

Tese de mestrado, Bioinformática e Biologia Computacional, Universidade de Lisboa, Faculdade de Ciências, 2019Em 2016, a Agência Internacional de Pesquisa sobre o Cancro da Organização Mundial de Saúde lançou a primeira base de dados de biomarcadores de exposição, chamada Exposome-Explorer. Para construir a base de dados, mais de 8500 citações foram manualmente analisadas, mas apenas 480 foram consideradas relevantes e usadas para extrair informação para integrar a base de dados. Curar manualmente uma base de dados é uma tarefa demorada e que requer especialistas capazes de recolher e analisar dados que se encontram espalhados por milhões de artigos. Esta tese propõe o uso de técnicas de Recuperação de Informação com uma abordagem de aprendizagem supervisionada para classificar automaticamente artigos como relevantes ou irrelevantes para auxiliar o processo de criação e atualização da Exposome-Explorer. Esta abordagem restringe a literatura a um conjunto de publicações relevantes sobre biomarcadores de exposição de uma maneira eficiente, reduzindo o tempo e esforço necessários para identificar documentos relevantes. Além disso, as queries originais usadas pelos curadores para pesquisar sobre literatura de biomarcadores de exposição foram melhoradas para incluir alguns artigos relevantes que anteriormente não estavam a ser encontrados. Os dados manualmente recolhidos d a Exposome-Explorer, foram usados para treinar e testar os modelos de aprendizagem automática (classificadores). Vários parâmetros e seis algoritmos diferentes foram avaliados para averiguar quais previam melhor a relevância de um artigo com base no título, resumo ou metadados. O melhor classificador foi construído com o algoritmo SVM e treinado com os resumos dos artigos, obtendo um recall de 85.8%.Este classificador reduz o número de citações sobre biomarcadores dietéticos a serem manualmente analisadas pelos curadores em quase 88%,classificando apenas incorrectamente 14.2% dos artigos relevantes.Esta metodologia também pode ser aplicada a outros dados de biomarcadores ou ser adaptada para auxiliar o processo de criação manual de outras bases de dados químicas ou de doenças

New Advances in Melanoma

Melanoma is a very aggressive tumor which is derived from the transformation of pigment-producing cells termed the melanocytes. This cancer type accounts for most of the deaths associated with skin cancer as well as its incidence and is in constant evolution. Because of the rapid and very high metastatic potential of this tumor, melanoma prognosis has been quite poor for a long time. In the past decade, groundbreaking discoveries in the melanoma research field have led to the development of two main treatment strategies: combination therapies targeting specific kinases or combination therapies focused on immune checkpoint inhibitors (ICIs). These treatment approaches have become the standard of care in most cancer centers and significantly improved the prognosis and overall survival of advanced melanoma patients. Nevertheless, many patients do not benefit from or even respond to these treatments. It is therefore essential to better comprehend the phenomenon of drug resistance, immune escape mechanisms, as well as to search for alternative treatment strategies. In addition, strong predictive biomarkers are desperately needed to improve clinical efficacy. The aim of this Special Issue is to present recent advances in the field of melanoma research, in which the abovementioned areas represent the primary focus, and other relevant themes are also discussed

MALDI imaging mass spectrometry in clinical proteomics research of gastric cancer tissues

In the presented thesis, matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry was used for the proteomic analysis of gastric cancer tissue samples, with the aims of 1) identifying proteins that predict disease outcome of patients with intestinal-type gastric cancer after surgical resection, and 2) generating a proteomic classifier that determines HER2-status in order to aid in therapy decision with regard to trastuzumab (Herceptin) administration. In the first study, a seven-protein signature was found to be associated with an unfavorable overall survival independent of major clinical covariates after analyzing 63 intestinal-type primary resected gastric cancer samples by MALDI imaging. Of these seven proteins, three could be identified as CRIP1, HNP-1, and S100-A6, and validated immunohistochemically on tissue microarrays of an independent validation cohort (n=118). While HNP-1 and S100-A6 were found to further subdivide early (UICC-I) and late stage (UICC-II-III) patients into different prognostic groups, CRIP1, a protein previously unknown in gastric cancer, was confirmed as a novel and independent prognostic factor for all patients in the validation cohort. The protein pattern described here serves as a new independent indicator of patient survival complementing the previously known clinical parameters in terms of prognostic relevance. In the second study, we hypothesized that MALDI imaging mass spectrometry may be useful for generating a classifier that may determine HER2-status in gastric cancer. This assumption was based on previous results where HER2-status could be reliably predicted in breast cancer patients. Here, 59 gastric cryo tissue samples were analyzed by MALDI imaging and the obtained proteomic profiles were used to create HER2 prediction models using different classification algorithms. Astonishingly, the breast cancer proteomic classifier from the previous study was able to correctly predict HER2-status in gastric cancers with a sensitivity of 65% and a specificity of 92%. In order to create a universal classifier for HER2-status, breast and non-breast cancer samples were combined, which increased sensitivity to 78%; specificity was 88%. This study provides evidence that HER2-status can be identified on a proteomic level across different cancer types suggesting that HER2 overexpression may constitute a widely spread molecular event independent of the tumor entity.Im Rahmen dieser Doktorarbeit wurden zwei Arbeiten publiziert, in denen die bildgebende Massenspektrometrie als zentrale Methode zur proteomischen Analyse von Magenkarzinomgeweben eingesetzt wurde. Dabei wurden folgende Ziele verfolgt: 1. die Identifizierung prognostischer Proteinmarker für Patienten mit intestinalem Magenkarzinom, und 2. die Generierung eines proteomischen Klassifikators zur Bestimmung des HER2-Status zur Entscheidungshilfe für eine Behandlung mit Trastuzumab (Herzeptin). In der ersten Studie wurde eine Signatur bestehend aus sieben Proteinsignalen gefunden, deren Überexpression unabhängig von anderen klinischen Parametern ein schlechtes Gesamtüberleben der Patienten indizieren. Hierzu wurden 63 Gewebeproben von Patienten mit Magenkarzinom intestinalen Typs mittels MALDI Imaging analysiert. Drei der sieben Proteinsignale konnten als CRIP1, HNP-1 und S100-A6 identifiziert werden. Diese wurden anschließend an einem unabhängigen Patientenkollektiv (n=118) immunhistochemisch anhand von Tissue Microarrays validiert. Dabei zeigte sich, dass die beiden Proteine HNP-1 und S100-A6 bestehende klinische Gruppen nach ihrem Risiko weiter aufstratifizieren konnten; HNP-1 Magenkarzinompatienten im frühen Stadium (UICC I) und S100-A6 Patienten im fortgeschrittenen Stadium (UICC II-III). Darüber hinaus konnte CRIP1 als unabhängiger prognostischer Faktor für alle Patienten des Validierungskollektives bestätigt werden. Perspektivisch könnte die hier beschriebene Proteinsignatur vorhandene klinische Parameter als neuer und unabhängiger Indikator für das Überleben von Magenkrebspatienten ergänzen. In der zweiten Studie wurden Proteinexpressionsmuster benutzt, um den HER2-Status in Magenkrebsgeweben vorauszusagen; denn seit kurzem ist der epidermale Wachstumsfaktor-Rezeptor HER2 eine wichtige tumorbiologische Zielstruktur bei der Behandlung von Magenkrebspatienten mit dem therapeutischen Antikörper Trastuzumab. In einer vorherigen Studie konnten wir die Machbarkeit der HER2-Status-Bestimmung durch MALDI Imaging erfolgreich anhand von Brustkrebsproben demonstrieren. Unter der Annahme, dass der HER2-Überexpression – unabhängig vom Tumortyp – charakteristische molekulare Veränderungen zugrunde liegen, wurde untersucht, ob eine Bestimmung des HER2-Status in Magenkrebspatienten mit Hilfe von Proteinexpressionsmustern aus Brustkrebspatienten erfolgen kann. Hierzu wurden, zusätzlich zu den bereits vorhandenen 48 Brustkrebsgeweben, 59 Magenkrebsfälle mittels MALDI Imaging analysiert und verschiedene HER2-Klassifikationsmodelle erstellt und verglichen. Der HER2-Status in Magenkrebsfällen konnte mit einem Mammakarzinom-spezifischen Profil mit einer Sensitivität von 65% und einer Spezifität von 92% bestimmt werden. Zusätzlich wurden die Expressionsprofile aller vorhandenen Tumorarten zusammengeführt, um einen universellen HER2-Klassifikator zu erstellen. Dies führte zu einer verbesserten Vorhersagequalität (Sensitivität: 78%, Spezifität: 88%). Dass sich der HER2-Status über verschiedene Tumorentitäten hinweg auf proteomischer Ebene bestimmen lässt, legt nahe, dass die Überexpression von HER2 ein unabhängiges molekulares Ereignis darstellt, ungeachtet der Herkunft des Tumors. Zudem unterstreichen die Ergebnisse das diagnostische Potential der bildgebenden Massenspektrometrie zur schnellen und zuverlässigen Bestimmung von tumorbiologischen Zielstrukturen, wie HER2

Methods for the evaluation of biomarkers in patients with kidney and liver diseases: multicentre research programme including ELUCIDATE RCT

Background: Protein biomarkers with associations with the activity and outcomes of diseases are being identified by modern proteomic technologies. They may be simple, accessible, cheap and safe tests that can inform diagnosis, prognosis, treatment selection, monitoring of disease activity and therapy and may substitute for complex, invasive and expensive tests. However, their potential is not yet being realised. Design and methods: The study consisted of three workstreams to create a framework for research: workstream 1, methodology – to define current practice and explore methodology innovations for biomarkers for monitoring disease; workstream 2, clinical translation – to create a framework of research practice, high-quality samples and related clinical data to evaluate the validity and clinical utility of protein biomarkers; and workstream 3, the ELF to Uncover Cirrhosis as an Indication for Diagnosis and Action for Treatable Event (ELUCIDATE) randomised controlled trial (RCT) – an exemplar RCT of an established test, the ADVIA Centaur® Enhanced Liver Fibrosis (ELF) test (Siemens Healthcare Diagnostics Ltd, Camberley, UK) [consisting of a panel of three markers – (1) serum hyaluronic acid, (2) amino-terminal propeptide of type III procollagen and (3) tissue inhibitor of metalloproteinase 1], for liver cirrhosis to determine its impact on diagnostic timing and the management of cirrhosis and the process of care and improving outcomes. Results: The methodology workstream evaluated the quality of recommendations for using prostate-specific antigen to monitor patients, systematically reviewed RCTs of monitoring strategies and reviewed the monitoring biomarker literature and how monitoring can have an impact on outcomes. Simulation studies were conducted to evaluate monitoring and improve the merits of health care. The monitoring biomarker literature is modest and robust conclusions are infrequent. We recommend improvements in research practice. Patients strongly endorsed the need for robust and conclusive research in this area. The clinical translation workstream focused on analytical and clinical validity. Cohorts were established for renal cell carcinoma (RCC) and renal transplantation (RT), with samples and patient data from multiple centres, as a rapid-access resource to evaluate the validity of biomarkers. Candidate biomarkers for RCC and RT were identified from the literature and their quality was evaluated and selected biomarkers were prioritised. The duration of follow-up was a limitation but biomarkers were identified that may be taken forward for clinical utility. In the third workstream, the ELUCIDATE trial registered 1303 patients and randomised 878 patients out of a target of 1000. The trial started late and recruited slowly initially but ultimately recruited with good statistical power to answer the key questions. ELF monitoring altered the patient process of care and may show benefits from the early introduction of interventions with further follow-up. The ELUCIDATE trial was an ‘exemplar’ trial that has demonstrated the challenges of evaluating biomarker strategies in ‘end-to-end’ RCTs and will inform future study designs. Conclusions: The limitations in the programme were principally that, during the collection and curation of the cohorts of patients with RCC and RT, the pace of discovery of new biomarkers in commercial and non-commercial research was slower than anticipated and so conclusive evaluations using the cohorts are few; however, access to the cohorts will be sustained for future new biomarkers. The ELUCIDATE trial was slow to start and recruit to, with a late surge of recruitment, and so final conclusions about the impact of the ELF test on long-term outcomes await further follow-up. The findings from the three workstreams were used to synthesise a strategy and framework for future biomarker evaluations incorporating innovations in study design, health economics and health informatics

Cell-Free Nucleic Acids

The deficits of mammography and the potential of noninvasive diagnostic testing using circulating miRNA profiles are presented in our first review article. Exosomes are important in the transfer of genetic information. The current knowledge on exosome-associated DNAs and on vesicle-associated DNAs and their role in pregnancy-related complications is presented in the next article. The major obstacle is the lack of a standardized technique for the isolation and measurement of exosomes. One review has summarized the latest results on cell-free nucleic acids in inflammatory bowel disease (IBD). Despite the extensive research, the etiology and exact pathogenesis are still unclear, although similarity to the cell-free ribonucleic acids (cfRNAs) observed in other autoimmune diseases seems to be relevant in IBD. Liquid biopsy is a useful tool for the differentiation of leiomyomas and sarcomas in the corpus uteri. One manuscript has collected the most important knowledge of mesenchymal uterine tumors and shows the benefits of noninvasive sampling. Microchimerism has also recently become a hot topic. It is discussed in the context of various forms of transplantation and transplantation-related advanced therapies, the available cell-free nucleic acid (cfNA) markers, and the detection platforms that have been introduced. Ovarian cancer is one of the leading serious malignancies among women, with a high incidence of mortality; the introduction of new noninvasive diagnostic markers could help in its early detection and treatment monitoring. Epigenetic regulation is very important during the development of diseases and drug resistance. Methylation changes are important signs during ovarian cancer development, and it seems that the CDH1 gene is a potential candidate for being a noninvasive biomarker in the diagnosis of ovarian cancer. Preeclampsia is a mysterious disease—despite intensive research, the exact details of its development are unknown. It seems that cell-free nucleic acids could serve as biomarkers for the early detection of this disease. Three research papers deal with the prenatal application of cfDNA. Copy number variants (CNVs) are important subjects for the study of human genome variations, as CNVs can contribute to population diversity and human genetic diseases. These are useful in NIPT as a source of population specific data. The reliability of NIPT depends on the accurate estimation of fetal fraction. Improvement in the success rate of in vitro fertilization (IVF) and embryo transfer (ET) is an important goal. The measurement of embryo-specific small noncoding RNAs in culture media could improve the efficiency of ET

Biomarkers and receptor expression in neuroendocrine tumours

Neuroendocrine tumours (NETs) are uncommon tumours which have a diverse biology. The aims of this thesis were to identify potential new biomarkers and further develop understanding of tumour biology in NETs. The following were assessed i) somatostatin receptor (SSTR) and dopamine-2 receptor (D2R) expression in NETs, ii)HER expression and their associated prognosis, iii) Angiopoietin expression in NETs and their prognostic significance, iv) proteomic analysis of serum and NET cell lines to identify novel markers and finally v) the role of 68Ga-DOTATATE PET in imaging of NETs. Immunohistochemical studies were performed to determine whether SSTR and D2R are co-expressed in NETs. D2R was co-expressed with SSTR-2 and -5 in 93% of low grade tumours, with lower co-expression in higher grade tumours. HER family of receptors are involved in oncogenesis; the expressions of these receptors were assessed. Immunohistochemical analysis of these receptors was performed in 82 cases. EGFR was expressed in 86%, HER-2 0%, HER-3 8.5% and HER-4 91.5%. The expression of EGFR was not associated with poor prognosis. Angiopoietins (Ang) are involved in tumourogenesis. Serum Ang-1 and Ang-2 were measured in patients and healthy controls. Ang-2 was significantly higher in patients compared to controls. Patients with Ang-2 levels >4756pg/ml had a shorter time to progression. Proteomic analysis using gel electrophoresis and LC/MS/MS of plasma from NET patients and established NET cell lines was performed to identify biomarkers. Proteomic cell line analysis identified 17 proteins in all cell lines including Mac-2 binding protein. We validated Mac-2 binding protein and it appears to be a potential marker for NETs. Finally, we performed a study to ascertain whether 68Ga-DOTATATE PET identifies more lesions in NET patients in whom 111In-DTPA-Octreotide showed faint/negative lesion uptake. 111In-DTPA-Octreotide scintigraphy identified 27 lesions compared to 168 lesions identified with Ga-68-DOTATATE PET. 68Ga-DOTATATE PET is a sensitive imaging modality for identifying NETs

Paradoxical Roles of Nanoparticles in Cancer Therapeutics and Carcinogenesis

Nanoparticles (NPs) are becoming increasingly common in consumer goods and are under investigation for a variety of industrial and biomedical applications. However, challenges in determining NP toxicity may prevent them from reaching their full potential. NPs cannot be treated as single class for toxicity evaluations. Even among particles made from the same material, particle-specific physical properties, including size, shape, surface charge, agglomeration state, and surface modifications have a strong effect on the toxicity. Even so, the obstacles to conclusively and reproducibly evaluating toxicity span all NP classes. NP literature is riddled with confusing and often contradictory reports regarding the biocompatibility of both engineered NPs, designed with biocompatibility as a priority, and NPs from occupational or environmental exposures. Incomplete NP characterization and sample inhomogeneity represent major confounding factors in disparate results from seemingly comparable study setups. Additionally, NPs can interfere with many conventional toxicity screening methods. Inappropriate doses, exposure routes, and toxicity endpoints further diminish the utility of many published studies.;Given the burgeoning interest in NP-based therapeutic agents, consistent, reliable standards are needed to ensure the biocompatibility of new formulations. To those ends, the synthesis, characterization, and in vitro toxicity of a multi-functional NP therapeutic were investigated (Chapter 2). Specifically, superparamagnetic iron oxide nanoparticles (SPIONs) were coated with amphiphilic polymer and functionalized with antisense oligonucleotides targeting survivin, an anti-apoptotic protein that is highly overexpressed in cancer. SPION physical properties, including particle size and composition, were characterized at each step of synthesis. Our results showed that the SPION platform is biocompatible and capable of delivering functional antisense oligonucleotides to regulate survivin expression; however, significant refinement of the DNA-to-SPION coupling step is needed. Applied clinically, antisense survivin coupled SPIONs can reduce the required dose of, adverse effects from, and resistance to, current cancer chemotherapy regimens.;In contrast to engineered NPs for biomedical applications, where real-world exposures would involve careful control of both exposure time- and dose, occupational NP exposures are variable, chronic, and difficult to model in laboratory settings. Chapter 3 focuses on identifying the mechanisms behind carbon nanotube (CNT)-induced malignant transformation of bronchial epithelial cells using a chronic in vitro exposure model. We specifically investigated the role of mesothelin (MSLN), a cell-surface protein that is highly overexpressed in many cancers, in the aggressive phenotype noted following chronic, low-dose CNT exposure. MSLN knockdown resulted in significantly decreased invasion, migration, colonies on soft agar, and tumor sphere formation. In vivo, MSLN knockdown cells formed smaller primary tumors and less metastases. The mechanism by which MSLN contributes to these more aggressive behaviors was investigated using Ingenuity Pathway Analysis, which predicted that increased MSLN could induce cyclin E, a cell cycle regulator known to be associated with human cancer. We found that MSLN knockdown cells had decreased cyclin E, and their proliferation rate was reverted to nearly that of untransformed cells. Cell cycle analysis results were consistent with the decreased rate of proliferation. Together, our results indicate a novel role of MSLN in the malignant transformation of bronchial epithelial cells following CNT exposure, suggesting its utility as a potential biomarker and drug target for CNT-induced malignancies.;As demonstrated by the two studies presented here, NPs have the potential to function as both cancer therapeutics and carcinogens. Careful evaluation of toxicity, ensuring that appropriate doses, assays, exposure routes, and endpoints are used, is imperative. Elucidating the physical properties and functionalization that contribute to toxicity, and the mechanisms of that toxicity, will allow NP benefits to be fully exploited while minimizing the risk of widespread, detrimental public health effects