Administració de fàrmacs dirigits i teràpia oncològica

L'ús de vehicles dirigits que inclouen conjugat el fàrmac actiu (nanoconjugados) permet una major eficàcia del tractament en la teràpia oncològica. A causa de la naturalesa xenobiótica i la toxicitat potencial de la majoria dels nanomaterials estudiats, les nanobiotecnologies emergents estan impulsant fortament el desenvolupament de nanoestructures basades en proteïnes autoassemblants que es caracteritzen per ser materials totalment biocompatibles. En aquest article, s'ha estudiat la combinació de pèptids dirigits a les cèl·lules tumorals i potenciadors de la fuita endosomal com a estratègia per augmentar l'eficàcia del nanoconjugat en la cèl·lula diana tumoral.El uso de vehículos dirigidos que incluyen conjugado el fármaco activo (nanoconjugados) permite una mayor eficacia del tratamiento en la terapia oncológica. Debido a la naturaleza xenobiótica y la toxicidad potencial de la mayoría de los nanomateriales estudiados, las nanobiotecnologías emergentes están impulsando fuertemente el desarrollo de nanoestructuras basadas en proteínas autoensamblables que se caracterizan por ser materiales totalmente biocompatibles. En este artículo, se ha estudiado la combinación de péptidos dirigidos a las células tumorales y potenciadores del escape endosomal como estrategia para aumentar la eficacia del nanoconjugado en la célula diana tumoral.Selective drug delivery (nanoconjugates generated by the cell-targeted drug and a nanoscale vehicle) could be a great asset in cancer treatment. Due to biosafety problems of the nanomaterials studied so far, emerging nanobiotechnologies are pushing the development of fully biocompatible nanosized protein materials based on the engineering of proteins that confer self-assembling at the nanoscale. This article studies the combined activity of a self-assembling protein and that of a pore- forming protein to improve its applicability as a drug carrier. This could result in a promising strategy to increase the effectiveness of nanoconjugates in the tumor target cell

Protein-based therapeutic killing for cancer therapies

Altres ajuts: CIBER-BBN (project NANOPROTHER) to A.V.; the Marató de TV3 foundation (TV32013-3930); CIBER-BBN (NanoMets 3) to R.M. in support of our research on cell-targeted antitumor drugs. N.S. by a predoctoral fellowship from the Gobierno de Navarra, R.D.O. by an overseas predoctoral fellowship from Conacyt (Gobierno de Méjico, 2016), U.U. received a Sara Borrell postdoctoral fellowship from ISCIII, and A.V. an Institución Catalana de Investigación y Estudios Avanzados (ICREA) ACADEMIA award.The treatment of some high-incidence human diseases is based on therapeutic cell killing. In cancer this is mainly achieved by chemical drugs that are systemically administered to reach effective toxic doses. As an innovative alternative, cytotoxic proteins identified in nature can be adapted as precise therapeutic agents. For example, individual toxins and venom components, proapoptotic factors, and antimicrobial peptides from bacteria, animals, plants, and humans have been engineered as highly potent drugs. In addition to the intrinsic cytotoxic activities of these constructs, their biological fabrication by DNA recombination allows the recruitment, in single pharmacological entities, of diverse functions of clinical interest such as specific cell-surface receptor binding, self-activation, and self-assembling as nanoparticulate materials, with wide applicability in cell-targeted oncotherapy and theragnosis

Protein scaffolds in human clinics

Acord transformatiu CRUE-CSICFundamental clinical areas such as drug delivery and regenerative medicine require biocompatible materials as mechanically stable scaffolds or as nanoscale drug carriers. Among the wide set of emerging biomaterials, polypeptides offer enticing properties over alternative polymers, including full biocompatibility, biodegradability, precise interactivity, structural stability and conformational and functional versatility, all of them tunable by conventional protein engineering. However, proteins from non-human sources elicit immunotoxicities that might bottleneck further development and narrow their clinical applicability. In this context, selecting human proteins or developing humanized protein versions as building blocks is a strict demand to design non-immunogenic protein materials. We review here the expanding catalogue of human or humanized proteins tailored to execute different levels of scaffolding functions and how they can be engineered as self-assembling materials in form of oligomers, polymers or complex networks. In particular, we emphasize those that are under clinical development, revising their fields of applicability and how they have been adapted to offer, apart from mere mechanical support, highly refined functions and precise molecular interactions

Nanostructured antimicrobial peptides : The last push towards clinics

Altres ajuts: to EU COST Action CA 17140 and ICREA ACADEMIA awardPeptide drugs hold great potential for the treatment of infectious diseases due to their unconventional mechanisms of action, biocompatibility, biodegradability and ease of synthesis and modification. The increasing rising of bacterial strains resistant to classical antibiotics have pushed the development of new peptide-based antimicrobial therapies. In this context, over the past few years, different approaches have reached a clinical approval. Furthermore, the application of nanotechnological principles to the design of antimicrobial peptide-based composites increases even more the already known benefits of antimicrobial peptides as competent protein drugs. Then, we provide here an overview of the current strategies for antimicrobial peptide discovery and modification and the status of such peptides already under clinical development. In addition, we summarize the innovative formulation strategies for their application, focusing on the controlled self-assembly for the fabrication of antimicrobial nanostructures without the assistance of external nanocarriers, and with emphasis on bioengineering, design of ultra-short peptides and rising insights in bacterial selectivity

In Vivo Bactericidal Efficacy of GWH1 Antimicrobial Peptide Displayed on Protein Nanoparticles, a Potential Alternative to Antibiotics

Oligomerization of antimicrobial peptides into nanosized supramolecular complexes produced in biological systems (inclusion bodies and self-assembling nanoparticles) seems an appealing alternative to conventional antibiotics. In this work, the antimicrobial peptide, GWH1, was N-terminally fused to two different scaffold proteins, namely, GFP and IFN-γ for its bacterial production in the form of such recombinant protein complexes. Protein self-assembling as regular soluble protein nanoparticles was achieved in the case of GWH1-GFP, while oligomerization into bacterial inclusion bodies was reached in both constructions. Among all these types of therapeutic proteins, protein nanoparticles of GWH1-GFP showed the highest bactericidal effect in an in vitro assay against Escherichia coli, whereas non-oligomerized GWH1-GFP and GWH1-IFN-γ only displayed a moderate bactericidal activity. These results indicate that the biological activity of GWH1 is specifically enhanced in the form of regular multi-display configurations. Those in vitro observations were fully validated against a bacterial infection using a mouse mastitis model, in which the GWH1-GFP soluble nanoparticles were able to effectively reduce bacterial loads

Collaborative membrane activity and receptor-dependent tumor cell targeting for precise nanoparticle delivery in CXCR4+ colorectal cancer

Altres ajuts: EU COST Action CA 17140. AV received an ICREA ACADEMIA award.By the appropriate selection of functional peptides and proper accommodation sites, we have generated a set of multifunctional proteins that combine selectivity for CXCR4 cell binding and relevant endosomal escape capabilities linked to the viral peptide HA2. In particular, the construct T22-GFP-HA2-H6 forms nanoparticles that upon administration in mouse models of human, CXCR4 colorectal cancer, accumulates in primary tumor at levels significantly higher than the parental T22-GFP-H6 HA2-lacking version. The in vivo application of a CXCR4 antagonist has confirmed the prevalence of the CXCR4 tumor tissue selectivity over unspecific cell penetration, upon systemic administration of the material. Such specificity is combined with improved endosomal escape, what overall results in a precise and highly efficient tumor biodistribution. These data strongly support the functional recruitment as a convenient approach to generate protein materials for clinical applications. More precisely, they also support the unexpected concept that enhancing the unspecific membrane activity of a protein material does not necessarily compromise, but it can even improve, the selective cell targeting offered by an accompanying functional module. Statement of Significance: We have shown here that the combination of cell-penetrating and tumor cell-targeting peptides dramatically enhances precise tumor accumulation of protein-only nanoparticles intended for selective drug delivery, in mouse models of human colorectal cancer. This fact is a step forward for the rational design of multifunctional protein nanomaterials for improved cancer therapies

Controlling self-assembling and tumor cell-targeting of protein-only nanoparticles through modular protein engineering

Altres ajuts: EU COST Action CA 17140. Villaverde A received an ICREA ACADEMIA award. Unzueta was supported by PERIS program from the Health Department of la Generalitat de Catalunya.Modular protein engineering is suited to recruit complex and multiple functionalities in single-chain polypeptides. Although still unexplored in a systematic way, it is anticipated that the positioning of functional domains would impact and refine these activities, including the ability to organize as supramolecular entities and to generate multifunctional protein materials. To explore this concept, we have repositioned functional segments in the modular protein T22-GFP-H6 and characterized the resulting alternative fusions. In T22-GFP-H6, the combination of T22 and H6 promotes self-assembling as regular nanoparticles and selective binding and internalization of this material in CXCR4-overexpressing tumor cells, making them appealing as vehicles for selective drug delivery. The results show that the pleiotropic activities are dramatically affected in module-swapped constructs, proving the need of a carboxy terminal positioning of H6 for protein self-assembling, and the accommodation of T22 at the amino terminus as a requisite for CXCR4 cell binding and internalization. Furthermore, the failure of self-assembling as regular oligomers reduces cellular penetrability of the fusions while keeping the specificity of the T22-CXCR4 interaction. All these data instruct how multifunctional nanoscale protein carriers can be designed for smart, protein-driven drug delivery, not only for the treatment of CXCR4 human neoplasias, but also for the development of anti-HIV drugs and other pathologies in which CXCR4 is a relevant homing marker

Nanostructures for delivery of therapeutic oligonucleotides

Fluoropyrimidines, such as 5-fluorouracil (5-FU) and related prodrugs are considered one of the most-successful agents in the treatment of colorectal cancer, yet poor specificity and tumor cell resistance remain major limiting bottlenecks. In this work, we exploited the ability of two DNA nanostructures, DNA tetrahedron (Td) and rectangle DNA origami, to incorporate 5-fluoro-2¿-deoxyuridine (FdUn) oligomers. DNA nanostructures bearing FdUn revealed to be able to circumvent 5-FU low sensitivity of colorectal drug-resistant cancer cells. Both DNA nanostructures attained comparable cytotoxic effect yet Td displays higher antiproliferative action. DNA nanoscaffolds functionalized with FdUn exhibited an enhanced cytotoxicity and higher ability to trigger apoptosis in colorectal cancer cells relative to conventional 5-FU and FdU drugs, especially having cholesterol as internalization helper. In addition, we will describe the preparation of a protein nanoparticle carrying FdUn oligomers with high affinity for cancer stem cells preventing the formation of metastasis in mice. The present work shows that DNA and protein nanoparticles are privileged carriers for delivering fluoropyrimidines, opening new avenues to the development of promising therapeutics for cancer treatment

Lymphocyte infiltration and antitumoral effect promoted by cytotoxic inflammatory proteins formulated as self-assembling, protein-only nanoparticles

Two human proteins involved in the inflammatory cell death, namely Gasdermin D (GSDMD) and the Mixed Lineage Kinase Domain-Like (MLKL) protein have been engineered to accommodate an efficient ligand of the tumoral cell marker CXCR4, and a set of additional peptide agents that allow their spontaneous self-assembling. Upon production in bacterial cells and further purification, both proteins organized as stable nanoparticles of 46 and 54 nm respectively, that show, in this form, a moderate but dose-dependent cytotoxicity in cell culture. In vivo, and when administered in mouse models of colorectal cancer through repeated doses, the nanoscale forms of tumor-targeted GSDMD and, at a lesser extent, of MLKL promoted CD8+ and CD20+ lymphocyte infiltration in the tumor and an important reduction of tumor size, in absence of systemic toxicity. The potential of these novel pharmacological agents as anticancer drugs is discussed in the context of synergistic approaches to more effective cancer treatments