Low Molecular Weight Nucleoside Gelators: A Platform for Protein Aggregation Inhibition

© Copyright 2018 American Chemical Society. Low molecular weight nucleoside gelators hold great promise in drug delivery and particularly for the delivery of biologics because of their excellent biocompatibility. However, the influence of these gelators on protein aggregation inhibition has not yet been studied. Protein aggregation is the most significant cause of protein instability and can severely impact the biological activity of the protein, impairing the quality and safety of the formulation. Herein, we report the ability of a nucleoside-based gelator, N4-octanoyl-2′-deoxycytidine, to inhibit protein aggregation. Using turbidimetric, spectroscopic, and microscopic methods, we demonstrate that protein aggregation inhibition is dependent on gelator concentration. Moreover, we have found that the protein is still functionally active in the hydrogel

Supramolecular Nucleoside-Based Gel:Molecular Dynamics Simulation and Characterization of Its Nanoarchitecture and Self-Assembly Mechanism

Among the diversity of existing supramolecular hydrogels, nucleic acid-based hydrogels are of particular interest for potential drug delivery and tissue engineering applications because of their inherent biocompatibility. Hydrogel performance is directly related to the nanostructure and the self-assembly mechanism of the material, an aspect that is not well-understood for nucleic acid-based hydrogels in general and has not yet been explored for cytosine-based hydrogels in particular. Herein, we use a broad range of experimental characterization techniques along with molecular dynamics (MD) simulation to demonstrate the complementarity and applicability of both approaches for nucleic acid-based gelators in general and propose the self-assembly mechanism for a novel supramolecular gelator, N4-octanoyl-2′-deoxycytidine. The experimental data and the MD simulation are in complete agreement with each other and demonstrate the formation of a hydrophobic core within the fibrillar structures of these mainly water-containing materials. The characterization of the distinct duality of environments in this cytidine-based gel will form the basis for further encapsulation of both small hydrophobic drugs and biopharmaceuticals (proteins and nucleic acids) for drug delivery and tissue engineering applications

Surface directed modulation of supramolecular gel properties

Surface properties directly affect fibre architecture and stiffness of self-assembled cytidine based gel films.</p

Hydrophobicity of surface-immobilised molecules influences architectures formed via interfacial self-assembly of nucleoside-based gelators

Surface-mediated self-assembly has potential in biomaterial development but underlying rules governing surface-gelator interactions are poorly understood. Here, we correlate surface properties with structural characterization data of nucleoside-based gels obtained by GISAXS and GIWAXS and find that hydrophobicity descriptors (logP, polar surface area, aromaticity) are key predictors for the gel structures formed

Cellular and microenvironmental cues that promote macrophage fusion and foreign body response

During the foreign body response (FBR), macrophages fuse to form foreign body giant cells (FBGCs). Modulation of FBGC formation can prevent biomaterial degradation and loss of therapeutic efficacy. However, the microenvironmental cues that dictate FBGC formation are poorly understood with conflicting reports. Here, we identified molecular and cellular factors involved in driving FBGC formation in vitro. Macrophages demonstrated distinct fusion competencies dependent on monocyte differentiation. The transition from a proinflammatory to a reparative microenvironment, characterised by specific cytokine and growth factor programmes, accompanied FBGC formation. Toll-like receptor signalling licensed the formation of FBGCs containing more than 10 nuclei but was not essential for cell-cell fusion to occur. Moreover, the fibroblastmacrophage crosstalk influenced FBGC development, with the fibroblast secretome inducing macrophages to secrete more PDGF, which enhanced large FBGC formation. These findings advance our understanding as to how a specific and timely combination of cellular and microenvironmental factors is required for an effective FBR, with monocyte differentiation and fibroblasts being key players

Development of a nanocapsule-loaded hydrogel for drug delivery for intraperitoneal administration

Intraperitoneal (IP) drug delivery of chemotherapeutic agents, administered through hyperthermal intraperitoneal chemotherapy (HIPEC) and pressurized intraperitoneal aerosolized chemotherapy (PIPAC), is effective for the treatment of peritoneal malignancies. However, these therapeutic interventions are cumbersome in terms of surgical practice and are often associated with the formation of peritoneal adhesions, due to the catheters inserted into the peritoneal cavity during these procedures. Hence, there is a need for the development of drug delivery systems that can be administered into the peritoneal cavity. In this study, we have developed a nanocapsule (NCs)-loaded hydrogel for drug delivery in the peritoneal cavity. The hydrogel has been developed using poly(ethylene glycol) (PEG) and thiol-maleimide chemistry. NCs-loaded hydrogels were characterized by rheology and their resistance to dilution and drug release were determined in vitro. Using IVIS® to measure individual organ and recovered gel fluorescence intensity, an in vivo imaging study was performed and demonstrated that NCs incorporated in the PEG gel were retained in the IP cavity for 24 h after IP administration. NCs-loaded PEG gels could find potential applications as biodegradable, drug delivery systems that could be implanted in the IP cavity, for example at a the tumour resection site to prevent recurrence of microscopic tumours

Mechanistic investigations into the encapsulation and release of small molecules and proteins from a supramolecular nucleoside gel in vitro and in vivo

Supramolecular gels have recently emerged as promising biomaterials for the delivery of a wide range of bioactive molecules, from small hydrophobic drugs to large biomolecules such as proteins. Although it has been demonstrated that each encapsulated molecule has a different release profile from the hydrogel, so far diffusion and steric impediment have been identified as the only mechanisms for the release of molecules from supramolecular gels. Erosion of a supramolecular gel has not yet been reported to contribute to the release profiles of encapsulated molecules. Here, we use a novel nucleoside-based supramolecular gel as a drug delivery system for proteins with different properties and a hydrophobic dye and describe for the first time how these materials interact, encapsulate and eventually release bioactive molecules through an erosion-based process. Through fluorescence microscopy and spectroscopy as well as Small Angle X-ray scattering, we show that the encapsulated molecules directly interact with the hydrogel fibres - rather than being physically entrapped in the gel network. The ability of these materials to protect proteins against enzymatic degradation is also demonstrated here for the first time. In addition, the released proteins were proven to be functional in vitro. Real-time fluorescence microscopy together with macroscopic release studies confirm that erosion is the key release mechanism. In vivo, the gel completely degrades after two weeks and no signs of inflammation are detected, demonstrating its in vivo safety. By establishing the contribution of erosion as a key driving force behind the release of bioactive molecules from supramolecular gels, this work provides mechanistic insight into the way molecules with different properties are encapsulated and released from a nucleoside-based supramolecular gel and sets the basis for the design of more tailored supramolecular gels for drug delivery applications

Repair of bone defects in vivo using tissue engineered hypertrophic cartilage grafts produced from nasal chondrocytes

The regeneration of large bone defects remains clinically challenging. The aim of our study was to use a rat model to use nasal chondrocytes to engineer a hypertrophic cartilage tissue which could be remodelled into bone in vivo by endochondral ossification. Primary adult rat nasal chondrocytes were isolated from the nasal septum, the cell numbers expanded in monolayer culture and the cells cultured in vitro on polyglycolic acid scaffolds in chondrogenic medium for culture periods of 5–10 weeks. Hypertrophic differentiation was assessed by determining the temporal expression of key marker genes and proteins involved in hypertrophic cartilage formation. The temporal changes in the genes measured reflected the temporal changes observed in the growth plate. Collagen II gene expression increased 6 fold by day 7 and was then significantly downregulated from day 14 onwards. Conversely, collagen X gene expression was detectable by day 14 and increased 100-fold by day 35. The temporal increase in collagen X expression was mirrored by increases in alkaline phosphatase gene expression which also was detectable by day 14 with a 30-fold increase in gene expression by day 35. Histological and immunohistochemical analysis of the engineered constructs showed increased chondrocyte cell volume (31–45 μm), deposition of collagen X in the extracellular matrix and expression of alkaline phosphatase activity. However, no cartilage mineralisation was observed in in vitro culture of up to 10 weeks. On subcutaneous implantation of the hypertrophic engineered constructs, the grafts became vascularised, cartilage mineralisation occurred and loss of the proteoglycan in the matrix was observed. Implantation of the hypertrophic engineered constructs into a rat cranial defect resulted in angiogenesis, mineralisation and remodelling of the cartilage tissue into bone. Micro-CT analysis indicated that defects which received the engineered hypertrophic constructs showed 38.48% in bone volume compared to 7.01% in the control defects. Development of tissue engineered hypertrophic cartilage to use as a bone graft substitute is an exciting development in regenerative medicine. This is a proof of principal study demonstrating the potential of nasal chondrocytes to engineer hypertrophic cartilage which will remodel into bone on in vivo transplantation. This approach to making engineered hypertrophic cartilage grafts could form the basis of a new potential future clinical treatment for maxillofacial reconstruction

Surface-Mediated Supramolecular Self-Assembly of Protein, Peptide, and Nucleoside Derivatives: From Surface Design to the Underlying Mechanism and Tailored Functions

Amongst the many parameters that have been explored to exercise control over self-assembly processes, the influence of surface properties on self-assembly has been recognized as important but has received considerably less attention than other factors. This is particularly true for biomolecule-derived self-assembling molecules such as protein, peptide or nucleobase derivatives. Due to their relevance for biomaterial and drug delivery applications, interest in these materials is increasing. As the formation of supramolecular structures from these biomolecule derivatives inevitably brings them into contact with surfaces of surrounding materials, understanding and control of the impact of the properties of these surfaces on the self-assembly process is important. In this review, we present an overview of the different surface parameters that have been used and studied for the direction of the self-assembly of protein, peptide and nucleoside-based molecules. The current mechanistic understanding of these processes will be discussed and potential applications of surface-mediated self-assembly will be outlined