Lysine Dendrimers Research Articles

Self-Assembled Nanoparticles from the Amphiphilic Prodrug of Resiquimod for Improved Cancer Immunotherapy.

Tumor-associated macrophages (TAMs) usually adopt a tumor-promoting M2-like phenotype, which largely impedes the immune response and therapeutic efficacy of solid tumors. Repolarizing TAMs from M2 to the antitumor M1 phenotype is crucial for reshaping the tumor immunosuppressive microenvironment (TIME). Herein, we developed self-assembled nanoparticles from the polymeric prodrug of resiquimod (R848) to reprogram the TIME for robust cancer immunotherapy. The polymeric prodrug was constructed by conjugating the R848 derivative to terminal amino groups of the linear dendritic polymer composed of linear poly(ethylene glycol) and lysine dendrimer. The amphiphilic prodrug self-assembled into nanoparticles (PLRS) of around 35 nm with a spherical morphology. PLRS nanoparticles could be internalized by antigen-presenting cells (APCs) in vitro and thus efficiently repolarized macrophages from M2 to M1 and facilitated the maturation of APCs. In addition, PLRS significantly inhibited tumor growth in the 4T1 orthotopic breast cancer model with much lower systemic side effects. Mechanistic studies suggested that PLRS significantly stimulated the TIME by repolarizing TAMs into the M1 phenotype and increased the infiltration of cytotoxic T cells into the tumor. This study provides an effective polymeric prodrug-based strategy to improve the therapeutic efficacy of R848 in cancer immunotherapy.

Bioorthogonal In Situ Polymerization of Dendritic Agents for Hijacking Lysosomes and Enhancing Antigen Presentation in Cancer Cells.

A low-generation lysine dendrimer, SPr-G2, responds to intracellular glutathione to initiate bioorthogonal in situ polymerization, resulting in the formation of large assemblies in mouse breast cancer cells. The intracellular large assemblies of SPr-G2 can interact with lysosomes to induce lysosome expansion and enhance lysosomal membrane permeabilization, leading to major histocompatibility complex class I upregulation on tumor cell surfaces and ultimately tumor cell death. Moreover, the use of the SPr-G2 dendrimer to conjugate the chemotherapeutic drug, camptothecin (CPT), can boost the therapeutic potency of CPT. Excellent antitumor effects in vitro and in vivo are obtained from the combinational treatment of the SPr-G2 dendrimer and CPT. This combinational effect also enhances antitumor immunity through promoting activation of cytotoxic T cells in tumor tissues and maturation of dendritic cells. This study can shed new light on the development of peptide dendritic agents for cancer therapy.

Enhanced rate of sarcosine N-carboxyanhydride polymerisation from a lysine dendrimer macroinitiator driven by neighbouring H-bonding effects

In situ FTIR monitoring revealed that sarcosine N-carboxyanhydride polymerisations were catalysed using a lysine dendrimer macroinitiator owing to hydrogen bonding interactions between the dendrimer core and monomer.

A Fluorophore-Labeled Lysine Dendrimer with an Oxo-Anion-Binding Motif for Tracking Gene Transfection.

A transfection vector based on a peptide dendrimer (1) has been developed and its abilities for DNA binding and transport have been investigated. By attaching a fluorophore to the vector system (1*), several steps in the transfection process could be monitored directly. As DLS and AFM studies showed, the labeled vector 1* condensed DNA into tightly packed aggregates able to enter eukaryotic cells. Co-localization experiments revealed that the ligand/plasmid complex is taken up by the endosomal pathway followed by an endosomal escape or lysosomal degradation. Afterwards, the plasmid DNA seems to enter the nucleus due to a breakdown of the nuclear envelope during mitosis, as only cells that have recently undergone mitosis showed H2B-GFP expression.

Histidylated poly(lysine) dendrimers for siRNA delivery

Since its discovery, small interfering RNA (siRNA) has revolutionised the field of gene silencing therapy because of its great potential in targeting diseases that were considered untreatable. Although several siRNA drugs are already in the market, many challenges remain to be addressed before siRNA therapies can realise their full potential including extra-hepatic delivery and improved endosomal escape. Dendritic polypeptides are attractive carriers for siRNA because of their wide range of defined structures that can be made. In this work, we report the on-going development of histidylated poly(lysine) dendrimers (G3KH) as carriers for siRNA. Histidine-capped lysine dendrimers were synthesised via HBTU/HOBt coupling reaction in solution. 1H NMR data confirms the structure of G3KH dendrimers; however, although traces of impurities were also observed. Histidylated lysine dendrimers could efficiently condense short double-strand DNA oligos (used as a model for siRNA) to form cationic nanoparticles of less than 100 nm in size

Polylysine dendrigraft is able to differentially impact Cutibacterium acnes strains preventing acneic skin.

With a view to reducing the impact of Cutibacteriumacnes (C.acnes) on acne vulgaris, it now appears interesting to modify the balance between acneic and non-acneic strains of C.acnes using moderate approach. In the present study, we identified that a G2 dendrigraft of lysine dendrimer (G2 dendrimer) was able to modify membrane fluidity and biofilm formation of a C.acnes acneic strain (RT5), whereas it appeared no or less active on a C.acnes non-acneic strain (RT6). Moreover, skin ex vivo data indicated that the G2 is able to decrease inflammation (IL1α and TLR-2) and improve skin desquamation after of C.acnes acneic strains colonization. Then, in vivo data confirmed, after C.acnes quantification by metagenomic analysis that the G2 cream after 28days of treatment was able to increase the diversity of C.acnes strains versus placebo cream. The data also showed a modification of the balance expression between C.acnes phylotype IA1 and phylotype II abundances. Taken together, the results confirm the interest of using soft compounds in cosmetic product for modifying phylotype abundances and diversity of C.acnes strains could be a new strategy for prevent acne vulgaris outbreak.

MD Simulation of AEDG Peptide Complexes with New K2R Dendrimer and Dendrigraft

Biocompatible peptide dendrimers and dendrigrafts have useful properties for application in biomedicine. In previous papers the computational approach for study lysine dendrimers and dendrigrafts as well as their complexes with various medical peptides was used. In this paper the comparison of complex formation between molecules of therapeutic AEDG tetrapeptide and novel K2R peptide dendrimer or DG2 dendrigraft of near the same size and charge was fulfilled. The systems consisting of 16 therapeutic AEDG tetrapeptide molecules and one dendrimer or one dendrigraft were studied by molecular dynamics simulation. Full atomic models of these molecules in water with explicit counterions were used for this goal. First of all, the process of complex formation was studied. It was obtained that peptide molecules were attracted by both branched molecules and were quickly adsorbed by them. Times of complexes formation as well as size, anisotropy and structure of each complex were calculated. It was demonstrated that both K2R dendrimer and DG2 dendrigraft are effective for complexation of these peptide molecules but new dendrimer complex is more stable than dendrigraft complex because it has almost twice more hydrogen bonds with peptide molecules and 33% more ion pairs with their charged groups.

Investigating the properties of l-lysine dendrimers through physico-chemical characterisation techniques and atomistic molecular dynamics simulations

Poly(l-lysine) dendrimers up to generation 6 have been analysed by SEC, Taylor dispersion analysis and atomistic molecular dynamics simulations to determine their properties, in what could be considered an understudied dendrimer series.

Bioinspired design of mannose-decorated globular lysine dendrimers promotes diabetic wound healing by orchestrating appropriate macrophage polarization

A large number of cytokines or growth factors have been used in the treatment of inflammation. However, they are highly dependent on an optimal delivery system with sufficient loading efficiency and protection of growth factors from proteolytic degradation. To develop the immunotherapy capacity of peptide dendrimers themselves, inspired by the structure and immunoregulatory functions of mannose-capped lipoarabinomannan (ManLAM), we thus propose a hypothesis that mannose-decorated globular lysine dendrimers (MGLDs) with precise molecular design can elicit anti-inflammatory activity through targeting and reprogramming macrophages to M2 phenotype. To achieve this, a series of mannose-decorated globular lysine dendrimers (MGLDs) was developed. Size-controlled MGLDs obtained were spherical with positive surface charges. The mean size ranged from 50-200 nm in varying generations and modification degrees. The initial screening study revealed that MGLDs have superior biocompatibility. When cocultured with MGLDs, mouse bone marrow-derived macrophages (BMDMs) acquired an anti-inflammatory M2 phenotype characterized by significant mannose receptor (MR) clustering on the cell surface and the elongated shape, an increased production of transforming growth factor (TGF)-β1, interleukin (IL)-4 and IL-10, a downregulated secretory of IL-1β, IL-6, and tumor necrosis factor (TNF)-α, and increased ability to induce fibroblast proliferation. Then in vivo studies further demonstrated that topical administration of optimized MGLDs accelerates wound repair of full-thickness cutaneous defects in type 2 diabetic mice via M2 macrophage polarization. Mechanistically, MGLDs treatment showed an enhanced closure rate, collagen deposition, and angiogenesis, along with mitigated inflammation modulated by a suppressed secretory of pro-inflammation cytokines, and increased production of TGF-β1. These findings provide the first evidence that the bioinspired design of MGLDs can direct M2 macrophage polarization, which may be beneficial in the therapy of injuries and inflammation.

Design and characterization of an in vivo injectable hydrogel with effervescently generated porosity for regenerative medicine applications

Injectable hydrogels that polymerize directly in vivo hold significant promises in clinical settings to support the repair of damaged or failing tissues. Existing systems that allow cellular and tissue ingrowth after injection are limited because of deficient porosity and lack of oxygen and nutrient diffusion inside the hydrogels. Here is reported for the first time an in vivo injectable hydrogel in which the porosity does not pre-exist but is formed concomitantly with its in situ injection by a controlled effervescent reaction. The hydrogel tailorable crosslinking, through the reaction of polyethylene glycol with lysine dendrimers, allows the mixing and injection of precursor solutions from a dual-chamber syringe while entrapping effervescently generated CO2 bubbles to form highly interconnected porous networks. The resulting structures allow preserving modular mechanical properties (from 12.7 ± 0.9 to 29.9 ± 1.7 kPa) while being cytocompatible and conducive to swift cellular attachment, proliferation, in-depth infiltration and extracellular matrix deposition. Most importantly, the subcutaneously injected porous hydrogels are biocompatible, undergo tissue remodeling and support extensive neovascularisation, which is of significant advantage for the clinical repair of damaged tissues. Thus, the porosity and injectability of the described effervescent hydrogels, together with their biocompatibility and versatility of mechanical properties, open broad perspectives for various regenerative medicine or material applications, since effervescence could be combined with a variety of other systems of swift crosslinking. Statement of significanceA major challenge in hydrogel design is the synthesis of injectable formulations allowing easy handling and dispensing in the site of interest. However, the lack of adequate porosity inside hydrogels prevent cellular entry and, therefore, vascularization and tissue ingrowth, limiting the regenerative potential of a vast majority of injectable hydrogels. We describe here the development of an acellular hydrogel that can be injected directly in situ while allowing the simultaneous formation of porosity. Such hydrogel would facilitate handling through injection while providing a porous structure supporting vascularization and tissue ingrowth.

Synthesis, characterization and topical application of novel bifunctional peptide metallodendrimer

Present research work focuses on the synthesis of lower generation bifunctional peptide metallodendrimer followed by its characterization using IR, Mass and NMR (1H and 13C) and finally exploring it for treating acne and wounds. Metallodendrimer was formulated into Tamarind Seed Polysaccharide (TSP) based hydrogel at 7% w/w concentration and analyzed for its properties of spreadability and bioadhesion. Testing of in-vitro antibacterial activity, sterility, in-vivo wound healing efficacy and stability studies followed the formulation development stage. The explored peptide metallodendrimer is a complex of second generation lysine dendrimers and zinc, exhibiting antibacterial activity against Propionibacterium acne, Staphylococcus epidermidis, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. In-vivo wound healing studies in comparison with marketed product indicated significant performance by metallodendrimer with respect to percent re-epithelization and wound contracture. The results were supported by concurrent improvement in various histopathological parameters such as cellularity, granulation, epithelization, vascular density, collagen staining intensity with less number of average mast cells and absence of scar formation. A bifunctional peptide metallodendrimer based platform technology was fabricated and duly evaluated for potential activities.

Bactericidal Activity of Non-Cytotoxic Cationic Nanoparticles against Clinically and Environmentally Relevant Pseudomonas spp. Isolates.

Difficult-to-treat bacterial infections caused by resistant human and plant pathogens severely afflict hospitals, and concern the agri-food sectors. Bacteria from the Pseudomonadaceae family, such as P. aeruginosa, P. putida, P. fluorescens, and P. straminea, can be responsible for severe nosocomial infections in humans. P. fragi is the major cause of dairy and meat spoilage, while P. syringae can infect a wide range of economically important plant species, including tobacco, kiwi, and tomato. Therefore, a cationic water-soluble lysine dendrimer (G5-PDK) was tested on several species of Pseudomonas genus. Interestingly, G5-PDK demonstrated variable minimum inhibitory concentrations (MICs), depending on their pigment production, on Pseudomonas aeruginosa (1.6-> 6.4 µM), MICs = 3.2–6.4 µM on P. putida clinical isolates producing pyoverdine, and very low MICs (0.2–1.6 µM) on strains that produced non-pigmented colonies. Time-kill experiments established the rapid bactericidal activity of G5-PDK. In the cytotoxicity experiments on human keratinocytes, after 4 h of treatment with G5-PDK at concentrations 16–500 × MIC, more than 80% of viable cells were observed, and after 24 h, the selectivity indices were maintained above the maximum value reported as acceptable. Due to its proven bactericidal potency and low cytotoxicity, G5-PDK should be seriously considered to counteract clinically and environmentally relevant Pseudomonas isolates.

Computing Zagreb and Randić Indices of PEG-Cored Dendrimers Used for Drug and Gene Delivery

Zagreb and Randić indices are the most commonly used degree-based topological indices in the study of drug design and development. In molecular topology, M-polynomials are also used to calculate the degree-based topological indices of chemical structures. In this paper, we derive the M-polynomials for the PEG-cored PAMAM, carbosilane, and poly (lysine) dendrimers and calculate their first, second, and second modified Zagreb indices and the Randić index.

Molecular Dynamics and Spin-Lattice NMR Relaxation in $$\alpha$$- and $$\varepsilon$$-Polylysine

The NMR relaxation method is widely used in various biomedical applications. Investigation of model homopeptides is an important step for understanding of structure and NMR properties of more complex branched peptides based on lysine monomers, for example, dendrimers, dendrigrafts and dendritic polymer brushes. In this paper, we perform molecular dynamics simulation of two linear lysine peptides with the same number of lysine monomers but with different connection between them through $$\alpha$$ - or $$\varepsilon$$ -peptide bonds. We obtained that the end-to-end distance and radius of gyration are smaller and radial density near the center of mass is essentially larger and decrease faster with radial distance for $$\alpha$$ -lysine peptide than for $$\varepsilon$$ -lysine peptide. Orientational mobility of $$\hbox {CH}_2$$ groups in both peptides could be described by second order orientational autocorrelation function and by spin-lattice NMR relaxation time. We calculated both functions and found that the relaxation of vector in side chains of $$\alpha$$ -lysine peptide is slightly faster in comparison with mobility of $$\hbox {CH}_2$$ groups in main chain of $$\varepsilon$$ -lysine peptide. Thus the big difference between the relaxation rates of these two types of $$\hbox {CH}_2$$ groups in lysine dendrimers obtained recently both in NMR and in simulation is mainly due to dendrimer effect and not due to difference in position (side or main chain) of $$\hbox {CH}_2$$ group in short linear lysine fragments. This result allows to use NMR for discrimination between $$\alpha$$ -lysine and $$\varepsilon$$ -lysine peptides as well as between linear lysine peptides (or their mixtures) and lysine dendrimers.

Synthesis and Characterization of Dendrimer-Based Polysarcosine Star Polymers: Well-Defined, Versatile Platforms Designed for Drug-Delivery Applications.

This paper describes the synthesis of star polymers designed for future drug-delivery applications. A generation-5 lysine dendrimer was used as a macroinitiator for the ring-opening polymerization of the sarcosine N-carboxyanhydride monomer to produce 32-arm star polymers with narrow molar mass distributions and desirable hydrodynamic size control. Fluorescent dye-labeled polymers were dosed in mice to measure plasma pharmacokinetics. Long circulation times were observed, representing ideal properties for biophysical targeting of tumors. In vivo efficacy of one of these star polymers conjugated to the therapeutic molecule SN-38 was evaluated in mice bearing SW620 xenografted tumors to demonstrate high antitumor activity and low body weight loss compared to the SN-38 prodrug irinotecan and this shows the potential of these delivery systems. As a further build, we demonstrated that these star polymers can be easily chain-end-functionalized with useful chemical moieties, giving opportunities for future receptor-targeting strategies. Finally, we describe the synthetic advantages of these star polymers that make them attractive from a pharmaceutical manufacturing perspective and report characterization of the polymers with a variety of techniques.

Interaction of lysine dendrimer with 8 and 16 molecules of EDR peptide

Dendrimers are frequently used for drug molecules delivery to different cells or organs. In our previous papers we used computer simulation to study the complex formation between dendrimers and dendrigrafts with different short regulatory peptides. The goal of present paper is to study interaction and the possibility of complex formation between lysine dendrimer and molecules of therapeutic EDR peptide. The system consisting of one lysine dendrimer of the second generation and 8 or 16 therapeutic EDR peptide molecules in water with explicit counterions was studied by computer simulation. The method of molecular dynamics and full atomic model were used for this goal. It was obtained that EDR peptide molecules become adsorbed by lysine dendrimer and form stable complex with it. Structure and conformational properties of this complex were studied. It was demonstrated that formation of complex occurs mainly due to electrostatic interaction between oppositely charged dendrimer and peptide molecules. Such complexes could be used in future for delivery of these or similar peptide molecules to the targeted tissues and organs.

Poly(lysine) Dendrimers Form Complexes with siRNA and Provide Its Efficient Uptake by Myeloid Cells: Model Studies for Therapeutic Nucleic Acid Delivery.

The disruption of the cellular pathways of protein biosynthesis through the mechanism of RNA interference has been recognized as a tool of great diagnostic and therapeutic significance. However, in order to fully exploit the potential of this phenomenon, efficient and safe carriers capable of overcoming extra- and intracellular barriers and delivering siRNA to the target cells are needed. Recently, attention has focused on the possibility of the application of multifunctional nanoparticles, dendrimers, as potential delivery devices for siRNA. The aim of the present work was to evaluate the formation of dendriplexes using novel poly(lysine) dendrimers (containing lysine and arginine or histidine residues in their structure), and to verify the hypothesis that the use of these polymers may allow an efficient method of siRNA transfer into the cells in vitro to be obtained. The fluorescence polarization studies, as well as zeta potential and hydrodynamic diameter measurements were used to characterize the dendrimer:siRNA complexes. The cytotoxicity of dendrimers and dendriplexes was evaluated with the resazurin-based assay. Using the flow cytometry technique, the efficiency of siRNA transport to the myeloid cells was determined. This approach allowed us to determine the properties and optimal molar ratios of dendrimer:siRNA complexes, as well as to demonstrate that poly(lysine) dendrimers may serve as efficient carriers of genetic material, being much more effective than the commercially available transfection agent Lipofectamine 2000. This outcome provides the basis for further research on the application of poly(lysine) dendrimers as carriers for nucleic acids in the field of gene therapy.

Bottlebrush-like highly efficient antibacterial coating constructed using α-helical peptide dendritic polymers on the poly(styrene-b-(ethylene-co-butylene)-b-styrene) surface.

Infectious diseases induced by pathogenic bacteria are the major causes for the failure of medical implants. Meanwhile, the drug-resistance is steadily developed because of the large and even inappropriate use of antibiotics. Therefore, the development of antibacterial coating with non-antibiotic-based agents on the surfaces of medical implants and devices has been an urgent need. Herein, we propose a bottlebrush-like antibacterial coating on a poly(styrene-b-(ethylene-co-butylene)-b-styrene) (SEBS) triblock copolymer surface by UV-induced graft polymerization of poly(ethylene glycol) (PEG) acrylate terminated poly(lysine dendrimer). This PEG-conjugated antibacterial polymer possessed a substructure of α-helical backbone and cation dendrimer side chains stretching in the radial directions of the helix. The introduction of lysine peptide dendrimers endowed the prepared antibacterial polymer with precisely controlled characteristics of its local cation density, amphipathic composition as well as three-dimensional (3D) conformation to improve interactions with bacterial membranes. The antimicrobial assay and biocompatibility assay results showed that 96.83% of S. aureus and 99.99% of E. coli were killed after being in contact with the antibacterial coating, while no toxicity to mammalian cells or no hemolysis was detected. This antimicrobial activity was further confirmed by the molecular dynamics simulation results, which demonstrated that the employment of lysine peptide dendrimers enhanced the electrostatic interaction and hydrogen bonding between the brush and bacterial membranes remarkably. Such bottlebrush-like antibacterial coating constructed using α-helical peptide dendritic polymers may become an effective strategy for manufacturing antibacterial products for biomedical uses.

Investigation of complex of lysine dendrimer of 2nd generation with molecules of therapeutic KED peptide by computer simulation

Lysine dendrimers could be used in many biomedical applications. For example these dendrimers could be used for delivery of short regulatory peptides consisting of several aminoacid residues. We investigated earlier interactions between lysine dendrigraft of 2nd generation and molecules of KED peptide. In present paper we study interaction of lysine dendrimer and molecules of KED peptide. The system containing one dendrimer of 2nd generation and 8 molecules of KED in water with explicit account of counterions was studied by computer simulation. The method of molecular dynamics was used for this goal. We obtained that formation of complex consisting of the dendrimer and all peptide molecules occurs during initial time (t < 40 ns) of simulation. The size, anisotropy of shape and radial density profile of stable complex (after t > 40ns) were studied also. We have shown that formation of complex occurs due to electrostatic interaction between oppositely charged dendrimer. At the same time other interactions, for example hydrogen bonds, also give their contribution to this process. Stable dendrimer-peptide complex has size close to 1,5nm and small shape anisotropy. Density of dendrimer atoms is highest in the centre of complex while density of peptides atoms has maximum at radial distance r=1nm. It total we have shown that lysine dendrimers is suitable carrier for molecules of KED peptide.

Lysine-based dendrimer with double arginine residues.

Due to their well-defined structure, multivalency, biocompatibility, and low toxicity, lysine dendrimers can be used as safe and efficient nanocarriers for drug and gene delivery. One useful strategy for improving the gene delivery properties of dendrimers is modification with arginine amino acid (Arg) residues. Incorporation of Arg residues could be favorable for the enhancement in transfection efficiency of lysine based dendrimers. In this work, we have synthesized a new second-generation poly-l-lysine dendrimer with repeating units containing two linear Arg residues between neighboring lysine branching points (Lys-2Arg dendrimer) and studied its physicochemical properties. We confirmed the structure of Lys-2Arg dendrimer using various one- and two-dimensional 1H and 13C NMR spectroscopy methods. Comparison of T1H relaxation data for Lys-2Arg and Lys-2Lys dendrimers showed that the replacement of double Lys residues with double Arg residues resulted in a sharp decrease in the mobility of methylene groups in side segments and in the main chain of ε-Lys inner segments. We suggest that this unexpected effect is caused by a guanidine–guanidine pairing effect in water, which leads to entanglements between dendrimer branches.