Data analysis for discovering the protein profile dynamics of the human ovarian follicular fluid and BRAF mutated metastatic melanoma tissue. : -

Abstract: Proteomics is widely utilized to understand the function of cellular processes at the molecular level. Using liquid chromatography interfaced with mass spectrometry (LC-MS)-based proteomics, thousands of proteins can be identified and quantified in a single experiment and their relationship and interactions can be analyzed. This makes the analysis of high-throughput proteomics data a cornerstone in the escalating field of translational medicine. Our group has been conducting deep mining LC-MS-based proteomics studies on two complex medical conditions that affect a high rate of the world population, female infertility and malignant melanoma (MM). To study female reproductive disorders, our group profiled the protein composition of the ovarian follicular fluid (FF) since it constitutes the microenvironment in which the oocyte develops during antral stages until follicular rupture at ovulation. In addition, it is believed that the FF mirrors what happens at the molecular level in the ovary and plasma due to pathological disorders. In the case of MM, we profiled the protein composition of metastatic tumor tissue from patients with BRAF mutation. The large amount of data generated from these experiments involves challenges related to data processing, analysis, and visualization of the results. The main challenge in complex disease pathology is the unraveling of the data from experimental outputs. In most cases the answer lies within that biological sample – the challenge is to analyze it and understand the meaning of the data.In this thesis, I performed data analyses to interrogate proteomics data (high resolution LC-MS expression data sets) from a bioinformatics and biostatistical point of view. Using different workflows, analyses and mathematical principles, I combined biological knowledge with bioinformatics and biostatistical approaches to integrate proteomics, clinical, and histopathological data in order to obtain new relevant biological insights from protein profiles of ovarian follicular fluids and MM tissues.The strategy applied in paper I, allowed us to describe progressive proteomic changes occurring in the FF during the ovulation process linked with oocyte maturation, hormone regulation and release of the oocyte. Here, we studied the most detailed temporal ovulatory interval, which included five time points. Paper II constituted the first large-scale proteomic characterization of FF extracted from small antral follicles (SAF) (6.1±0.4 mm) in their natural state. Using a multivariate approach, a signature of proteins appeared to play a role in oocyte maturation and oocyte meiotic resumption already from the early follicular stage. As a follow-up, paper III reported for the first time evidence of proteomic alterations occurring in the FF of SAF of polycystic ovaries (PCO). Alterations were associated with the dysfunction of follicular growth and subsequent oocyte competence usually observed in PCO syndrome. Furthermore, uncharacterized or poorly characterized proteins identified in the FF of unstimulated SAF were assessed and their functionality during folliculogenesis was described in paper IV (manuscript). In paper V, data analysis revealed for the first time that the high expression, in the MM tumor, of the B-raf V600E (mutated) protein could be a significant risk factor for poorer prognosis of patients with stages 3 or 4 of MM. A follow-up of this finding was performed on a larger cohort of patients with BRAF mutation, in which subgroups of patients with different mortality risks were identified and associated with the activation of different BRAF-related pathways, such as the immune response.Supported by data-driven results, this thesis characterized the protein profile dynamics of human ovarian FF during folliculogenesis (paper I-IV) and malignant melanoma tissue of patients with BRAF mutation (paper V). Findings from paper I to IV may open up new pathways for augmenting or attenuating subsequent oocyte viability in the pre-ovulatory follicle when it is ready to undergo ovulation, which may be of importance to future advances in reproductive medicine. On the other hand, findings from paper V may enable the eventual delineation of patient response therapy for MM with BRAF mutation.