Human fluids pretreatment using three-phase partitioning systems for the diagnosis and prognosis of lung cancer

• Autor(a) principal: Mendes, Maria Silvina Marques
• Data de Publicação: 2021
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10773/31941
• topic: Lung cancer; Pentraxin-3; Depletion; Aqueous biphasic systems; Three-phase partitioning systems

Lung cancer is responsible for the highest rate of cancer mortality worldwide. The 5-year survival rate of lung cancer is less than 15%, mainly because most lung cancer patients are diagnosed at late stages. Therefore, early detection of lung cancer represents one of the most effective approaches to help reduce the high associated mortality and morbidity. Technological advances in genomics and proteomics fields have disclosed many novel cancer biomarkers, which can be used in early cancer detection. Pentraxin-3 is one of these promising biomarkers, found in serum, for lung cancer diagnosis and prognosis. However, human serum is a complex matrix whose protein profile mainly comprises human serum albumin (HSA) and immunoglobulin G (IgG). These proteins can interfere or even mask the detection and quantification of less abundant ones, such as PTX3, leading to false results. To contribute towards an earlier and more reliable diagnosis of lung cancer, there is a need to implement sample pretreatment steps that permit the depletion of the most abundant proteins and the concentration of tumor biomarkers of interest. In this context, three-phase partitioning (TPP) based on aqueous biphasic systems (ABS) are an alternative to conventional pretreatment techniques that use expensive affinity resins or volatile organic solvents. The main goal of this work is to develop new TPP systems based on ABS composed of polymers/copolymers and salts as alternative serum pretreatment strategies. To this end, novel TPP-ABS formed by the homopolymers PPG 400, PEG 400, PEG 600, PEG 1000 and PEG 2000, and the copolymers Pluronic PE6200, Pluronic PE6400, Pluronic L35 and UCON, and a citrate buffered salt, were investigated in terms of their performance to simultaneous deplete HSA and IgG and extract the biomarker PTX-3 to the polymer-rich phase. According to the results obtained, TPP systems formed by PPG 400, PEG 1000, PEG 2000 and UCON allow depletion efficiencies above 80% for both proteins, with the systems formed by PEG 1000 and UCON allowing depletions efficiencies reaching 100% for both abundant proteins. Among the best studied systems, only the TPP system formed by PEG 1000 allows the complete extraction of PTX-3 towards the polymer-rich phase, while keeping the complete depletion of IgG and HSA. ELISA assays demonstrate the better performance of this TPP foe the PTX-3 quantification in the pretreated human serum samples, thus representing a promising alternative for the pretreatment of human serum for the diagnosis and prognosis of lung cancer.