Lipid Nanoparticles with Targeting Capabilities
Authorship
A.O.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
A.O.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
Defense date
01.30.2026 09:00
01.30.2026 09:00
Summary
Lipid nanoparticles (LNPs) are a clinically validated platform for nucleic acid delivery, yet their lack of inherent cell-type specificity remains a major limitation for targeted cancer therapy. Conventional LNP formulations tend to undergo broad cellular internalization via generic endocytic pathways, resulting in uptake by both cancerous and non-cancerous cells. This study explored a biomimetic strategy to modulate LNP-cell interactions through the incorporation of membrane proteins derived from lung cancer cells, with the objective of assessing whether such modification could influence nanoparticle selectivity without compromising physicochemical stability or biocompatibility. Lipid Nanoparticles formulated with the SC8 ionizable lipid were used as the nanoparticle platform due to their favorable stability and delivery characteristics. Membrane proteins extracted from A549 human lung adenocarcinoma cells were incorporated into SC8 LNPs using two functionalization strategies: in situ incorporation during nanoparticle assembly (pre-formulation) and adsorption onto pre-formed nanoparticles (post-formulation). These approaches were compared to evaluate their effects on LNP properties, membrane protein retention, cellular uptake, and cytotoxicity. Physicochemical characterization using dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA) demonstrated that all formulations formed stable, nanoscale particles with PDIs within the range suitable for cellular uptake. Pre-formulation membrane protein incorporation resulted in slightly increased size heterogeneity and more pronounced surface charge alterations, whereas post-formulation modification preserved a more uniform nanoparticle profile. Particle concentration measurements confirmed that membrane protein functionalization did not adversely affect nanoparticle yield or colloidal stability. Quantitative and qualitative protein analysis confirmed the successful association of membrane proteins with SC8 LNPs using both strategies, with higher protein retention observed for post-formulation modification. All formulations exhibited strong and quantifiable intrinsic fluorescence, enabling reliable comparison in subsequent uptake studies. Cellular uptake was evaluated in A549 lung cancer cells and MRC-5 normal lung fibroblasts. In A549 cells, LNP uptake was largely preserved across all formulations, with only a modest enhancement observed for post-formulation membrane-functionalized LNPs at prolonged incubation. In contrast, pre and post-membrane protein functionalization produced a statistically significant but small reduction in uptake by MRC-5 fibroblasts at specific time points. This indicates partial modulation of non-specific internalization rather than robust exclusion of off-target cells. The magnitude of this effect was limited and did not constitute strong active targeting. Cytotoxicity assessment using the MTT assay demonstrated that both unmodified and membrane-functionalized SC8 LNPs were well tolerated, with no significant reduction in cell viability observed. Overall, these results demonstrate that membrane protein functionalization of SC8 LNPs is technically feasible and preserves nanoparticle stability and biocompatibility. Although robust homotypic targeting was not achieved, the ability to selectively modulate non-specific uptake by normal fibroblasts demonstrates that biomimetic surface engineering can meaningfully influence cellular uptake. This work provides a mechanistic foundation for the further optimization of membrane protein-based targeting strategies for nanotherapeutic applications.
Lipid nanoparticles (LNPs) are a clinically validated platform for nucleic acid delivery, yet their lack of inherent cell-type specificity remains a major limitation for targeted cancer therapy. Conventional LNP formulations tend to undergo broad cellular internalization via generic endocytic pathways, resulting in uptake by both cancerous and non-cancerous cells. This study explored a biomimetic strategy to modulate LNP-cell interactions through the incorporation of membrane proteins derived from lung cancer cells, with the objective of assessing whether such modification could influence nanoparticle selectivity without compromising physicochemical stability or biocompatibility. Lipid Nanoparticles formulated with the SC8 ionizable lipid were used as the nanoparticle platform due to their favorable stability and delivery characteristics. Membrane proteins extracted from A549 human lung adenocarcinoma cells were incorporated into SC8 LNPs using two functionalization strategies: in situ incorporation during nanoparticle assembly (pre-formulation) and adsorption onto pre-formed nanoparticles (post-formulation). These approaches were compared to evaluate their effects on LNP properties, membrane protein retention, cellular uptake, and cytotoxicity. Physicochemical characterization using dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA) demonstrated that all formulations formed stable, nanoscale particles with PDIs within the range suitable for cellular uptake. Pre-formulation membrane protein incorporation resulted in slightly increased size heterogeneity and more pronounced surface charge alterations, whereas post-formulation modification preserved a more uniform nanoparticle profile. Particle concentration measurements confirmed that membrane protein functionalization did not adversely affect nanoparticle yield or colloidal stability. Quantitative and qualitative protein analysis confirmed the successful association of membrane proteins with SC8 LNPs using both strategies, with higher protein retention observed for post-formulation modification. All formulations exhibited strong and quantifiable intrinsic fluorescence, enabling reliable comparison in subsequent uptake studies. Cellular uptake was evaluated in A549 lung cancer cells and MRC-5 normal lung fibroblasts. In A549 cells, LNP uptake was largely preserved across all formulations, with only a modest enhancement observed for post-formulation membrane-functionalized LNPs at prolonged incubation. In contrast, pre and post-membrane protein functionalization produced a statistically significant but small reduction in uptake by MRC-5 fibroblasts at specific time points. This indicates partial modulation of non-specific internalization rather than robust exclusion of off-target cells. The magnitude of this effect was limited and did not constitute strong active targeting. Cytotoxicity assessment using the MTT assay demonstrated that both unmodified and membrane-functionalized SC8 LNPs were well tolerated, with no significant reduction in cell viability observed. Overall, these results demonstrate that membrane protein functionalization of SC8 LNPs is technically feasible and preserves nanoparticle stability and biocompatibility. Although robust homotypic targeting was not achieved, the ability to selectively modulate non-specific uptake by normal fibroblasts demonstrates that biomimetic surface engineering can meaningfully influence cellular uptake. This work provides a mechanistic foundation for the further optimization of membrane protein-based targeting strategies for nanotherapeutic applications.
Direction
DEL PINO GONZALEZ DE LA HIGUERA, PABLO ALFONSO (Tutorships)
POLO TOBAJAS, ESTER (Co-tutorships)
DEL PINO GONZALEZ DE LA HIGUERA, PABLO ALFONSO (Tutorships)
POLO TOBAJAS, ESTER (Co-tutorships)
Court
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Ruthenium cumulenic intermediates for bioorthogonal catalysis
Authorship
L.P.P.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
L.P.P.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
Defense date
01.30.2026 09:00
01.30.2026 09:00
Summary
The development of new bioorthogonal reactions has revolutionized the areas of synthetic and biological chemistry, given their high potential for applications in fields such as biomedicine. In this context, transition metal complexes can be powerful catalysts due to the mechanistic versatility they offer. Specifically, the introduction of species such as metallic vinylidenes and allenylidenes in bioorthogonal chemistry has not yet been achieved, but it could be of interest to transfer new reactivities into this field. In this Master dissertation, we present the development of a ruthenium-catalyzed synthesis of a furan derivative from a 3-butyne-1,2-diol precursor in aqueous environments. Mechanistic experimental studies have allowed us to propose that the reaction takes place through ruthenium vinylidene and allenylidene intermediate species. Moreover, we have performed preliminary experiments for its translation to more complex biological media. In particular, we have demonstrated the feasibility of this reaction at low concentrations, mild temperatures and in aqueous media. Besides, we have investigated its tolerance towards biomolecules and biologically relevant media. Moreover, we have preliminary tested the viability of this transformation inside living mammalian cells (HeLa cells) and bacteria (S. aureus and B. thuringiensis), observing its potential for furan synthesis within biological environments. Finally, aiming to further investigate this type of reactivity in vivo, we have synthesized a new probe for the synthesis of a fluorescent furan, which would allow for the monitoring of the reaction in biological contexts (e.g., in cellulo) by fluorescence microscopy.
The development of new bioorthogonal reactions has revolutionized the areas of synthetic and biological chemistry, given their high potential for applications in fields such as biomedicine. In this context, transition metal complexes can be powerful catalysts due to the mechanistic versatility they offer. Specifically, the introduction of species such as metallic vinylidenes and allenylidenes in bioorthogonal chemistry has not yet been achieved, but it could be of interest to transfer new reactivities into this field. In this Master dissertation, we present the development of a ruthenium-catalyzed synthesis of a furan derivative from a 3-butyne-1,2-diol precursor in aqueous environments. Mechanistic experimental studies have allowed us to propose that the reaction takes place through ruthenium vinylidene and allenylidene intermediate species. Moreover, we have performed preliminary experiments for its translation to more complex biological media. In particular, we have demonstrated the feasibility of this reaction at low concentrations, mild temperatures and in aqueous media. Besides, we have investigated its tolerance towards biomolecules and biologically relevant media. Moreover, we have preliminary tested the viability of this transformation inside living mammalian cells (HeLa cells) and bacteria (S. aureus and B. thuringiensis), observing its potential for furan synthesis within biological environments. Finally, aiming to further investigate this type of reactivity in vivo, we have synthesized a new probe for the synthesis of a fluorescent furan, which would allow for the monitoring of the reaction in biological contexts (e.g., in cellulo) by fluorescence microscopy.
Direction
Mascareñas Cid, Jose Luis (Tutorships)
Mascareñas Cid, Jose Luis (Tutorships)
Court
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Autocatalytic self-replication of peptide amphiphiles through native chemical ligation
Authorship
B.O.T.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
B.O.T.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
Defense date
01.30.2026 09:00
01.30.2026 09:00
Summary
Peptide amphiphiles (PAs) are peptide-based molecules formed from the conjugation of a peptide sequence and a lipid tail. They can self-assemble into ordered nanostructures -such as micelles, vesicles and nanofibers- and can be generated by distinct ligation strategies. This project focuses on understanding the in situ synthesis of autocatalytic PAs via oxime chemistry an already well-studied ligation strategy- and transferring this knowledge to native chemical ligation (NCL), a biocompatible ligation strategy. This project was carried out at CiQUS in two stages, April to May 2025 and from September to December 2025. First, the reactivity of both chemistries and the in situ self-assembling capacity of the different sequences are discussed by a combination of spectroscopic, chromatographic and microscopic techniques, suggesting the big impact of the chemistry on the in situ autocatalytic PA formation. Lastly, the formation of 2 distinct PAs in NCL was studied -varying the reaction time and equivalents of the tail- showing how important the modulation of these 2 parameters is for the predominant PA specie. In conclusion, this study provides insight into the rationale design of PAs via a more biocompatible chemistry (NCL) for potential biomedical applications as the design of in situ synthesised and self-assembled PAs.
Peptide amphiphiles (PAs) are peptide-based molecules formed from the conjugation of a peptide sequence and a lipid tail. They can self-assemble into ordered nanostructures -such as micelles, vesicles and nanofibers- and can be generated by distinct ligation strategies. This project focuses on understanding the in situ synthesis of autocatalytic PAs via oxime chemistry an already well-studied ligation strategy- and transferring this knowledge to native chemical ligation (NCL), a biocompatible ligation strategy. This project was carried out at CiQUS in two stages, April to May 2025 and from September to December 2025. First, the reactivity of both chemistries and the in situ self-assembling capacity of the different sequences are discussed by a combination of spectroscopic, chromatographic and microscopic techniques, suggesting the big impact of the chemistry on the in situ autocatalytic PA formation. Lastly, the formation of 2 distinct PAs in NCL was studied -varying the reaction time and equivalents of the tail- showing how important the modulation of these 2 parameters is for the predominant PA specie. In conclusion, this study provides insight into the rationale design of PAs via a more biocompatible chemistry (NCL) for potential biomedical applications as the design of in situ synthesised and self-assembled PAs.
Direction
INSUA LOPEZ, IGNACIO (Tutorships)
INSUA LOPEZ, IGNACIO (Tutorships)
Court
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Semiconducting Covalent Organic Frameworks based on Trioxotriangulene Neutral Radicals
Authorship
S.T.P.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
S.T.P.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
Defense date
01.30.2026 09:00
01.30.2026 09:00
Summary
Over the past few years, covalent organic frameworks (COFs) have emerged as crystalline porous materials due to their well-defined structures, stable porosity, and chemical tuneability. These materials are constructed from organic building blocks connected through strong covalent bonds, and they can be rationally designed, making them highly interesting materials for catalysis, electronics and electrochemical energy storage, among other applications. However, their use in electronic and energy storage is often limited by their intrinsic low electrical conductivity. In this context, the incorporation of redox-active and conductive radical building blocks into COF structures has attracted growing attention as a potential solution to face this drawback. Trioxotriangulene (TOT) neutral radical derivatives are particularly promising for this purpose due to their high stability, multiple accessible redox states, and ability to delocalize unpaired electrons over extended pi-conjugated systems to promote high electrical conductivity. In this Master dissertation, we have synthesized and characterized two isoreticular radical-based COFs incorporating TOT building blocks, demonstrating that it is possible to retain their redox-active, conductive, and magnetic properties within the framework. The synthesized COFs were fully characterized by multiple spectroscopic techniques to elucidate their structure, chemical composition and physical properties. This work opens the way for the design of conductive COFs based on neutral organic radicals for applications in energy storage and spintronics.
Over the past few years, covalent organic frameworks (COFs) have emerged as crystalline porous materials due to their well-defined structures, stable porosity, and chemical tuneability. These materials are constructed from organic building blocks connected through strong covalent bonds, and they can be rationally designed, making them highly interesting materials for catalysis, electronics and electrochemical energy storage, among other applications. However, their use in electronic and energy storage is often limited by their intrinsic low electrical conductivity. In this context, the incorporation of redox-active and conductive radical building blocks into COF structures has attracted growing attention as a potential solution to face this drawback. Trioxotriangulene (TOT) neutral radical derivatives are particularly promising for this purpose due to their high stability, multiple accessible redox states, and ability to delocalize unpaired electrons over extended pi-conjugated systems to promote high electrical conductivity. In this Master dissertation, we have synthesized and characterized two isoreticular radical-based COFs incorporating TOT building blocks, demonstrating that it is possible to retain their redox-active, conductive, and magnetic properties within the framework. The synthesized COFs were fully characterized by multiple spectroscopic techniques to elucidate their structure, chemical composition and physical properties. This work opens the way for the design of conductive COFs based on neutral organic radicals for applications in energy storage and spintronics.
Direction
Souto Salom, Manuel (Tutorships)
Souto Salom, Manuel (Tutorships)
Court
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Light Modulated Properties of Self-Assembled Cyclic Peptide Nanotubes
Authorship
M.V.R.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
M.V.R.
Master's Degree in Chemistry at the Interface with Biology and Materials Science
Defense date
01.30.2026 09:00
01.30.2026 09:00
Summary
This work aims to develop new properties of supramolecular peptide nanotubes using light. Two different supramolecular systems were studied. In the first, a beta-unsaturated amino acid was incorporated into the structure of an amphipathic cyclopeptide. This allows for the control of the double bond isomerization using light, transitioning from a folded conformation that prevents self-assembly to a planar cyclopeptide geometry that favors nanotube formation. This approach aims to control the assembly/disassembly of these supramolecular structures, potentially allowing for the control of their biological activity. In the second supramolecular system, four cyclopeptides were anchored to a core with electrochemical activity. Controlling assembly with pH changes modulates this activity, opening the door for using light as a control method for these systems with potential energy storage capacity.
This work aims to develop new properties of supramolecular peptide nanotubes using light. Two different supramolecular systems were studied. In the first, a beta-unsaturated amino acid was incorporated into the structure of an amphipathic cyclopeptide. This allows for the control of the double bond isomerization using light, transitioning from a folded conformation that prevents self-assembly to a planar cyclopeptide geometry that favors nanotube formation. This approach aims to control the assembly/disassembly of these supramolecular structures, potentially allowing for the control of their biological activity. In the second supramolecular system, four cyclopeptides were anchored to a core with electrochemical activity. Controlling assembly with pH changes modulates this activity, opening the door for using light as a control method for these systems with potential energy storage capacity.
Direction
Granja Guillán, Juan Ramón (Tutorships)
Granja Guillán, Juan Ramón (Tutorships)
Court
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)
Pérez Meirás, María Dolores (Chairman)
POLO TOBAJAS, ESTER (Secretary)
LAZZARI , MASSIMO (Member)