Preparation of Dendrimeric antigen-silica particle composites

Abstract para Comunicación en el congreso tipo cartelThe desing and synthesis of new materials for biomedical applications is a high-priority research topic in a great number of biomedical areas. Moreover, advances in the fabrication of these materials are of growing interest in antibody-based diagnostic techniques. These materiasl consist in a solid support anchored with the desired bioactive molecules. Such solid supports need to be robust enough and posses surrounding reactive groups that enables the chemical bonding of the active components. Furthermore, these materials need to meet certain conditions to ensure biocompatibility and non-toxicity. In this sense, silica nanoparticles have been widely used. Our research involves the study of hybrid materials that combine the high functionality of silica nanoparticles with well defined size and controlled peripheral multivalence components like dendrimers. Dendrimer antigens, which are synthetic antigens where the role of the carrier protein is performed by a dendrimer, were supported on silica particles. These organic-inorganic hybrid materials were carefully characterized and the preparation methodology was confirmed to be highly reproducible. Such hybrid materials were used for the in vitro diagnosis of patien allergic to amoxicillin.1 Herein, we present the preparation of novel nano-materials containing new antigenics determinants of antibiotics. Amoxicillin, bencyl penicillin, clavulanic acid and its derivates were used to prepares diferent dendrimeric antigens supported on silca particles. These particles will be used to specifically and selectively detect and quantify IgE in sera from allergic patients. These new materials are a promising candidate to improve the practice of in vitro clinical diagnosis.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Modificación de aptámeros anti-gluten: efecto sobre sus características de enlace

En este trabajo de investigación se realizan estudios de truncado de los aptámeros descritos por el Grupo de Investigación de Electroanálisis de la Universidad de Oviedo para el reconocimiento de las proteínas del gluten (Gli 4 y Gli 1) y protegidos por la patente ES 2436861 B2. Se trata de mejorar tanto la afinidad del aptámero por la proteína como abaratar los costes de dicho aptámero

Silica Particle based nanocomposites for Specific IgE Determination to Betalactams

Current immunoassays for diagnosing betalactam (BL) allergy have limited sensitivity and poor specificity. These include ImmunoCAP, the commercial immunoassay, and the radioallergosorbent test (RAST), a handmade test performed only in specialized laboratories. Both are based on cellulose polymers modified with covalently attachedbetalactam-poly-L-lysine conjugates, which are able to interact with specific IgE (sIgE). In RAST, dendrimers have been used instead of poly-L-lysine in the formation of penicilloyl- conjugates, showing specific recognition by penicillin-sIgE. In this work, we propose the use of nanomaterials consisting on silica particles highly functionalized with drug-dendrimer conjugates (including mono- and bi-epitopic structures), as precise and controlled solid phases able to improve the tests’ reliability. The homogeneous composition of the nano-composites provides high reproducibility and quality, which is critical for hospital applications, while silica particles possess promising properties as biocompatibility, tuneable size, and functionalisation, and their preparation is affordable and reproducible. The use of nano-scaled silica particles improved the results of available immunoassays techniques in terms of sensitivity and specificity, providing the possibility of testing different betalactams, simultaneously.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Dendrímeros antigénicos sobre partículas de sílica para determinar IgE específicas de antibióticos betalactámicos

Los inmunoensayos empleados para el diagnóstico in vitro de alergias a antibióticos betalactámicos actualmente presentan una limitada sensibilidad y pobre especificidad, además de no estar disponibles de forma comercial para todos los antibióticos implicados en procesos alérgicos. En este trabajo, se han diseñado nanopartículas de sílica altamente funcionalizadas con dendrímeros y conjugadas con amoxicilina y bencilpenicilina con objeto de mejorar el diagnóstico clínico in vitro de alergia a estos antibióticos.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Dendrímeros Antigénicos fijados sobre nanopartículas para diagnostico in vitro de alergia a fármacos

Las Reacciones Adversas a Fármacos (RAF) constituyen actualmente un problema sanitario importante debido al gran desconocimiento sobre los mecanismos por los que se desencadenan, lo que se traduce en una ausencia de pruebas diagnósticas suficientemente sensibles y específicas para su diagnóstico. Este problema se ha manifestado en numerosos foros científicos, existiendo un acuerdo unánime entre todas las partes interesadas en este problema (academia, industria, asociaciones de pacientes) para el desarrollo de iniciativas encaminadas a resolver dichas limitaciones. Actualmente nos enfrentamos a dos grandes problemas a la hora de abordar el estudio de los aspectos relacionados con la respuesta inmune (los procesos de reacción adversa a antígenos hapténicos -proceso alérgico- y en particular a fármacos): i) menor sensibilidad de los métodos diagnósticos in vivo e in vitro que obliga a identificar las nuevas estructuras hapténicas que inducen las reacciones, y ii) una retirada de los kits comerciales habiendo dejado un vacío que debe ser solucionado. El grupo de investigación en el que se ha realizado este trabajo de tesis doctoral, ha desarrollado una investigación focalizada en dos objetivos básicos: i) disponer de una herramienta que nos permita estudiar el proceso de reconocimiento molecular antígeno-anticuerpo responsable de las reacciones alérgicas a fármacos; ii) desarrollar test in vitro para la detección de las reacciones alérgicas a fármacos. En base a todos los resultados prometedores obtenidos a lo largo de la trayectoria de investigación del grupo, en esta tesis se ha desarrollado una nueva línea de investigación. Esta se sustenta en el estudio de la síntesis de partículas de sílice como soporte sólido para los nano-conjugados desarrollados previamente dendrímero-hapteno, en los test in vitro utilizados actualmente para el diagnóstico de reacciones adversas a fármacos

Multiepitope Dendrimeric Antigen-Silica Particle Composites as Nano-Based Platforms for Specific Recognition of IgEs

β-lactam antibiotics (BLs) are the drugs most frequently involved in drug hypersensitivity reactions. However, current in vitro diagnostic tests have limited sensitivity, partly due to a poor understanding of in vivo drug-protein conjugates that both induce the reactions and are immunologically recognized. Dendrimeric Antigen-Silica particle composites (DeAn@SiO2), consisting on nanoparticles decorated with BL-DeAns are promising candidates for improving the in vitro clinical diagnostic practice. In this nano-inspired system biology, the synthetic dendrimer plays the role of the natural carrier protein, emulating its haptenation by drugs and amplifying the multivalence. Herein, we present the design and synthesis of new multivalent mono- and bi-epitope DeAn@SiO2, using amoxicillin and/or benzylpenicillin allergenic determinants as ligands. The homogeneous composition of nanoparticles provides high reproducibility and quality, which is critical for in vitro applications. The suitable functionalization of nanoparticles allows the anchoring of DeAn, minimizing the nonspecific interactions and facilitating the effective exposure to specific IgE; while the larger interaction area increments the likelihood of capturing specific IgE. This achievement is particularly important for improving sensitivity of current immunoassays since IgE levels in BL allergic patients are very low. Our data suggest that these new nano-based platforms provide a suitable tool for testing IgE recognition to more than one BL simultaneously. Immunochemical studies evidence that mono and bi-epitope DeAn@SiO2 composites could potentially allow the diagnosis of patients allergic to any of these drugs with a single test. These organic-inorganic hybrid materials represent the basis for the development of a single screening for BL-allergies.This work was supported by the Spanish Ministerio de Ciencia e Innovación (Proyectos de I+D+I «Programación Conjunta Internacional», EuroNanoMed 2019 (PCI2019-111825-2), Ministerio de Ciencia y Educación (PID2019-104293GB-I00), Instituto de Salud Carlos III (ISCIII) of MINECO (grants cofunded by ERDF: ‘‘Una manera de hacer Europa’’ (Euronanomed Program AC19/00082, PI20/01734, PI18/00095, RETIC ARADYAL, RD16/0006/0001 and RD16/0006/0012, and Miguel Servet II program (CPII20/00028)), Junta de Andalucía and Universidad de Málaga (UMA18-FEDERJA-007), Andalusian Regional Ministry of Health (PE-0172-2018) and Nicolas Monardes Program (RC-0004-2021).Ye