Absolute quantification of selenoproteins and selenometabolites in lung cancer human serum by column switching coupled to triple quadrupole inductively coupled plasma mass spectrometry

Abstract

Una de las causas más importantes de la alta tasa de mortalidad y la baja esperanza de vida del cáncer de pulmón es la detección en estadios avanzados. Por ello, existe una necesidad urgente de un diagnóstico precoz y de la búsqueda de nuevos biomarcadores selectivos. El selenio es un componente importante de las selenoproteínas y un potente antioxidante capaz de proteger frente al cáncer. En este trabajo se ha realizado por primera vez la cuantificación absoluta de selenio en selenoproteínas y el contenido total en selenometabolitos en suero de pacientes con cáncer de pulmón (CL) y controles sanos (HC). Para ello, se desarrolló un método de especiación simultánea de selenoproteínas mediante cromatografía de exclusión por tamaño (SEC) y cromatografía de afinidad (AF) con detección por ICP-QQQ-MS, y cuantificación por dilución isotópica (IDA) (SEC-AF-HPLC-SUID-ICP-QQQ-MS) para determinar la concentración de selenio en eGPx, SEPP1 y SeAlb, así como selenometabolitos totales, para encontrar alteraciones que puedan servir como biomarcadores de esta enfermedad. De la misma forma, se desarrolló un método basado en cromatografía de intercambio aniónico acoplado a ICP-QQQ-MS para cuantificar selenometabolitos (SeCys2, SeMeSeCys, SeMet, selenito y selenato) en las mismas muestras de suero.One of the most important causes of the high mortality rate and low life expectancy of lung cancer is the detection at advanced stages. Thus, there is an urgent need for early diagnosis and the search of new selective biomarkers. Selenium is an important constituent of selenoproteins and a powerful antioxidant able to protect against cancer. In this work, the absolute quantification of selenium in selenoproteins and the total content in selenometabolites has been performed for the first time in serum from lung cancer patients (LC) and healthy controls (HC). To this end, a method for the simultaneous speciation of selenoproteins using size exclusion chromatography (SEC) and affinity chromatography (AF) with detection by ICP-QQQ-MS, and quantification by isotopic dilution (IDA) (SEC-AF-HPLC-SUID-ICP-QQQ-MS) was developed to determine the selenium concentration in eGPx, SEPP1 and SeAlb, as well as total selenometabolites, to find alterations that may serve as biomarkers of this disease. In the same way, a method based on anion-exchange chromatography coupled to ICP-QQQ-MS was developed to quantify selenometabolites (SeCys2, SeMeSeCys, SeMet, selenite and selenate) in the same LC and HC serum samples. The results showed that the averaged concentrations of selenium in eGPx, SeAlb and selenite were significantly higher in LC patients (LC (eGPx: 21.24 ± 0.77 ng g−1; SeAlb: 49.56 ± 3.16 ng g−1 and Se(IV): 6.20 ± 1.22 ng g−1) than in HC group (eGPx: 16.96 ± 0.53 ng g−1; SeAlb: 38.33 ± 2.66 ng g−1 and Se(IV): 3.56 ± 0.55 ng g−1). In addition, the ratios between selenoproteins and selenometabolites have been calculated for the first to study their potential use as LC biomarkers. The rates eGPx/SEPP1, SEPP1/SeAlb, eGPx/Se(IV) and SEPP1/Se(IV) were significantly different between LC and HC groups.