Synthesis and applications in biomedicine of new dendrimeric structures

Specific cell interactions of surfaces grafted dendrimers have been applied to the study of cell adhesion. We have studied also the application of dendrimer-based uneven nanopatterns to evaluate the local RGD surface density effects on cell adhesion, as bioactive substrates to evaluate the impact of the RGD local surface density on the chondrogenic induction of adult human mesenchymal stem cells [1], [2], [3]. Dendrimers that are modified through their peripheral groups have been employed for many biomedical purposes. IgE detection via specific interactions with synthetic Dendrimeric Antigens (DeAn) is also one of the objectives that has been given more attention in our group. In this way, we have been developing different methodologies for the detection of immunoglobulins, as an in vitro method for the diagnosis of allergic reactions to drugs [4], [5]. PAMAM dendrimer has allowed us to develop useful methods for this purpose, for certain objectives we have required the development of new dendrimeric models. We have developed a new approach for the production of all-aliphatic polyamide dendrimers (BAPAD) by iterative 3,3′-diaminopivalic acid connections as building blocks for dendrimer construction. These dendrimers were studied in explicit solvent by atomistic forcefield-based molecular dynamics to characterize structural properties such as shape, radius and monomer distribution [6].Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Design and synthesis of new molecules based on indolium derivatives for two-photon absorption bioimaging applications

Bioimaging is the visualization of biological processes in real time, interfering as less as possible with these and using non-invasive methods. Among others, fluorescence methods have acquired a very important role in these purposes through the nature of the light. Bioimaging pretends to understand how our organism works identifing subcellular organelles, following cellular processes, quantifying ion or biological species and visualising interactions of molecules with their targets in living cells or tissues. In the last decades, two-photon (TP) microscopy is unseating classical one-photon (OP) microscopy due to its advantages, such as the use of lower energy excitation wavelengths or the possibility of focus in depth, among others. Nowadays, it is an interesting target design and develop of new probes for TP microscopy to biomaging. Fluorophores based on indolenines are a family of compounds with promising properties in this sense. In this work, we present the design, synthesis and characterization of new indolium derivatives with promising properties to be used in bioimaging applications in living cells with different purposes.Real Sociedad Española de Química, Jóvenes Invetigadores Químicos, Facultad de Ciencias Ambientales y Bioquímica (UCLM), Merck, Cortes de Castilla la Mancha, Reaxys (Elsevier), Diputación de Toledo, MestreLab Research, Sección Territorial de Castilla la Mancha (RSEQ), Grupo especializado en Nanociencia y Materiales Moleculares (RESEQ), Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Tuning pKa in new molecules based on indolines for two-photon absorption bioimaging applications

Fluorescence-based biosensors have become essential tools for modern biology, allowing real-time monitoring of biological processes within living cells. The differences between pH play a significant role in multiple biological process such as proliferation, apoptosis, endocytic process and defense. Therefore, intracellular fluorescent pH probes comprise one of the most widely used families of biosensors in microscopy. In this work, we present the design, synthesis and characterization of new one and two photon pH activatable fluorescent probes based on naphthalene-indoline derivatives with an off-on response to different pH environments. These compounds are easy to achieve with good yields and have good photophysical properties. Simple modifications in the structure of these compounds allow tuning their pKa easily and cover a width range of pH in their applications. The preliminary results obtained with these fluorescent probes indicate that they have promising properties to be use in bioimaging applications in living cells.Real Sociedad Española de Química y Grupo Especializado de Química Orgánica de la RSEQ•Universidade de Santiago de Compostela • CIQUS • Lilly • Mestrelab Research •Thieme •Scharlab • Acros • ABCR • AMSLab • GalChimia • Elsevier • The Journal of the American Chemical Society • ACS Omega • Organic Letters • The Journal of Organic Chemistry • Accounts of Chemical Research. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Design and synthesis of new dendrons for biomedical applications

Dendrons are excellent multifunctional platforms for many different applications. In particular, the development of multivalent molecular nanostructures, with a well-defined size and shape, has an enormous interest in the field of biomedicine1. Great efforts are being made in the design of new dendritic structures since their precise structure, multivalency and the possibility of functionalizing their terminal groups makes them a useful tool for certain applications. Recently, a new type of dendrimer (dendron) was designed and synthesized based on the iterative coupling of 2,2-Bis (AminoAlkyl) Propanamide units (BAPAD)2,3. This represents a versatile model when incorporating certain functionalities in its structure. However, the synthesis of these new dendritics structures was addressed using 3,3'-dichloropivalic acid as the starting substrate, which translated in the superficial amino groups of these new macromolecule being in relative positions 1,3. For certain applications, where the functionalization of the dendritics structures on its surface requires the introduction of relatively bulky groups, this may result in a problem of reactivity given the steric congestion. Here, we present the design and synthesis of new dendrons where the surface amino groups are in relative positions 1,7. Formally, it involves the preparation of dendrons with longer arms that can minimize the problems of steric congestion. These dendrons can be designed to modify surfaces superficially thanks to the versatility of the carboxylic acid which can be modified to introtoduce in the focal poin diferent functional groups (azido, amido, thiol, etc…).Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

New indolium fluorofores for two-photon absortion (2PA) bioimaging applications

The development of organic materials with 2PA has attracted intensive attention in the past two decades [1]. In two-photon bio-imaging applications the design of the chromophore requires to have a good cross-section (σ2PA) and good biological compatibility which depends on the molecular volume and polarity [2]. In this work, we present the design, synthesis and characterization of new indolium derivatives. These compounds are easy to achieve with good yields and good photophysical properties. In addition, time-dependent density functional theory (TDDFT) has been carried out to investigate the energy level of the ground and excited state. Their spectral properties and assays performed on cultured cells, demonstrate the potential of these compounds as fluorescent probes with application in two-photon bio-imaging.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Ministerio de Ciencia e Innovación-Spain (CTQ2013-41339-P and CTQ2015- 71896-REDT

Detecting hydroxyl radical with a new two-photon fluorescent probe in living cells

Reactive oxygen species (ROS) are known to be involved in the onset and development of multiple diseases, including cancer, cardiovascular diseases, neurodegenerative diseases and diabetes among others. Generation of ROS is a phenomenon that results from normal cell metabolism as well as from the response to certain pathologic stimuli like certain cytokines, xenobiotics and bacterial infection. These ROS are highly reactive, short-lived molecules that play critical roles in the living cell and it is well accepted that cellular oxidative stress results from the imbalance between generation and elimination of ROS in cells. In this work, we describe the design and synthesis of a two photon (TP) organelle-targeting activatable fluorescent probe. This probe is a naphthalene-indoline compound that targets specifically lysosomes, reaching these organelles in the fluorescence “off” state and minimizing background reactions. Upon arrival at the lysosomes, the probe is triggered and a fluorescence “on” signal is observed that can be combined with TP microscopy to image the lysosomal •OH in living cells.Real Sociedad Española de Química y Grupo Especializado de Química Orgánica de la RSEQ•Universidade de Santiago de Compostela • CIQUS • Lilly • Mestrelab Research •Thieme •Scharlab • Acros • ABCR • AMSLab • GalChimia • Elsevier • The Journal of the American Chemical Society • ACS Omega • Organic Letters • The Journal of Organic Chemistry • Accounts of Chemical Research. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Synthesis of fluorescent BAPAD dendrimeric structrures for biomedical applications

Comunicación oral presentada en el Congreso IV Encuentro sobre Dendrímeros, Santiago de Compostela, 2014In this work, we present the synthesis and characterization of BAPAD1 fluorescent Dendrimeric-Antigens (DeAn), to study the dendritic cell maturation as a test to detect drug allergy reactions. Recently our research group developed a new kind of dendrimer, called BAPAD1, that we have used in this work to obtain the dendrimeric moiety of the target molecule. To this avail we synthesized a generation two BAPAD dendrimer using cystamine as core. Then, the free amine groups on the surface of the dendrimer were functionalized with an amoxiciloyl group (AXO), the allergenic determinant to the beta-lactam antibiotic amoxicillin. By the reduction of the disulfur bond we obtained two dendrons with a thiol group in the focal point, useful to attach a fluorescent probe. We synthesize as fluorescent moiety a naphthalimide derivative with a maleimide group upon which the thiol group of the dendron is added by a click reaction.2 In this way we obtained the target molecule to be used in the basophil activation test. The fluorescent DeAn (F-DeAn) has been fully characterized by NMR and MS techniques, and their fluorescent properties well established in aqueous biological media. The fluorescent dendron without the haptenic moieties at the periphery has been also obtained and fully characterized as a control assay. Both molecules have been also characterized using molecular dynamics simulation calculations. We show also here how these dendrimeric structures interact with dendritic cells and are internalized by them.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Synthesis of fluorescent dendrimeric antigen efficiently internalized by human dendritic cells

A new fluorescent dendrimeric antigen (DeAn) based on a dendron with amoxicilloyl terminal groups has been synthetized. The synthesis implies a novel class of all-aliphatic polyamide dendrimer (BisAminoalkylPolyAmide Dendrimers, or BAPAD).[1] The introduction of a cystamine core allows the incorporation of this dendrons into a 1,8-naphthalimide fluorofore functionalized with a maleimide group. The fluorescence properties of this DeAn has been studied and compared with the properties of an equivalent dendron possessing amino-terminal groups. This DeAn has been used as a synthetic antigen in a biomedical assay that tests the amoxicillin sensitivity of dendritic cells (DC) from tolerant and allergic patients.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Preparation, photophysical properties and cell image of Naphthalimide-based fluorescent nanostructures for biomedical applications: BAPAD dendrimeric antigens and silica nanoparticles

We present here the synthesis and characterization of BAPAD [1] fluorescent Dendrimeric-Antigens (DeAn), to be applied in basophil activation as a test to detect drug allergy reactions. A second generation BAPAD dendrimer using cystamine as core that can be reducced to obtain two dendrons with a thiol group in the focal point, useful to attach a fluorescent probe, have been synthesized. The free amine groups on the surface of the dendrimer were functionalized with an amoxiciloyl group (AXO), the allergenic determinant to the beta-lactam antibiotic amoxicillin. The fluorescent DeAn (F-DeAn) has been fully characterized by NMR and MS techniques, and their fluorescent properties well established in aqueous biological media. F-DeAn without the haptenic moieties at the periphery has been also obtained and fully characterized as a control assay. Both molecules have been also characterized using molecular dynamics simulation calculations. We show here how these dendrimeric structures interacts with dendritic cells and are internalized by them. In a secon approximation, Naphthalimide-fluorscent silica nanoparticles have been prepared and characterized, to be used also in basophil activation test.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Diseño de dendrímeros amino-terminales basados en enlaces tipo amidas. Estructuras con menor repulsión estérica y síntesis a gran escala

La estructura ramificada y la elevada multivalencia que presentan los dendrímeros los convierten en excelentes compuestos con potenciales aplicaciones en el campo de la biomedicina. En concreto, los dendrímeros con grupos amino terminales han resultado ser una herramienta muy útil en este campo gracias a la gran versatilidad que presentan. Nuestro grupo de investigación tiene una gran experiencia en el diseño y síntesis de dendrímeros amino-terminales [1]. La inserción de estas unidades dendriméricas en complejos luminiscentes nos ha permitido obtener macromoléculas con excelentes propiedades como biomarcadores [2]. Además, la modificación superficial de implantes orales de titanio con este tipo de estructuras ha contribuido a mejorar la biocompatibilidad de los mismos [3]. En esta comunicación presentamos una nueva familia de dendrímeros de poliamida amino-terminales. Su diseño implica una mayor separación entre los grupos terminales, generando así estructuras con menor congestión estérica. La metodología sintética se basa en procedimientos bien establecidos, utilizando reactivos sencillos y económicos, con rendimientos superiores al 90%, en escala de gramos y evitando procedimientos de purificación tediosos [4