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| Author(s): |
Rebeca Kawahara
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Campinas, SP. |
| Institution: | Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia |
| Defense date: | 2015-04-28 |
| Examining board members: |
Adriana Franco Paes Leme;
Giuseppe Palmisano;
Inácio de Loiola Meirelles Junqueira de Azevedo;
Vitor Marcel Faça;
Marcio Chaim Bajgelman
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| Advisor: | Adriana Franco Paes Leme |
| Abstract | |
Mass spectrometry applied to proteomic analysis allows large-scale characterization and quantification of proteins in specific conditions or biological systems. In this thesis, we showed different mass spectrometry-based approaches for (1) the study of cell secreted or released from the cell-surface proteins (secretome) as a source of candidate biomarkers in cancer (2) validation of candidate biomarkers in human saliva of patients with oral cancer and (3) the study of the target substrates of an important metalloprotease involved in cancer progression, the ADAM17 (A disintegrin and metalloproteinase 17). Together with statistical and bioinformatics tools, such as clustering, heatmaps, network analysis and enrichment analysis, we found specific hyphothesis-driven proteins that were further validated using complementary quantitative and functional methods. In the context of cancer biomarker, a panel of candidate biomarkers was revealed from the secretome analysis of carcinoma, melanoma and non-tumorigenic cell lines. Among the carcinoma candidates, proteins involved in complement system were validated with higher expression in tissues (C3 and CFB) and saliva (C3, CFB, C4B and SERPINA1) from patients with oral squamous cell carcinoma. The function of agrin and perlecan, which were also found with increased expression in squamous cell carcinoma tissue, were studied in tumorigenic processes: migration, adhesion, proliferation and sensibility to cisplatin. In the context of ADAM17 degradome, mimecan, perlecan and glypican-1 (GPC1) were revealed as novel substrates and their cleavage sites were determined using mass spectrometry. Besides, the function and biological processes associated with GPC1 shedding were studied by identifying the binding partners of GPC1 in the extracellular media and by performing functional assays. In summary, mass spectrometry was present in all part of the scientific method, from hyphothesis generation using discovery proteomics to hyphothesis testing using targeted proteomics (AU) | |
| FAPESP's process: | 11/22421-2 - Determination of cleavage sites of recombinant ADAM-17 in human cells |
| Grantee: | Rebeca Kawahara Sakuma |
| Support Opportunities: | Scholarships in Brazil - Doctorate (Direct) |