3D-Printed Gastric Resident Electronics.

Long-term implantation of biomedical electronics into the human body enables advanced diagnostic and therapeutic functionalities. However, most long-term resident electronics devices require invasive procedures for implantation as well as a specialized receiver for communication. Here, a gastric resident electronic (GRE) system that leverages the anatomical space offered by the gastric environment to enable residence of an orally delivered platform of such devices within the human body is presented. The GRE is capable of directly interfacing with portable consumer personal electronics through Bluetooth, a widely adopted wireless protocol. In contrast to the passive day-long gastric residence achieved with prior ingestible electronics, advancement in multimaterial prototyping enables the GRE to reside in the hostile gastric environment for a maximum of 36 d and maintain ≈15 d of wireless electronics communications as evidenced by the studies in a porcine model. Indeed, the synergistic integration of reconfigurable gastric-residence structure, drug release modules, and wireless electronics could ultimately enable the next-generation remote diagnostic and automated therapeutic strategies.

Biocompatibility of hydrogel-based scaffolds for tissue engineering applications.

Recently, understanding of the extracellular matrix (ECM) has expanded rapidly due to the accessibility of cellular and molecular techniques and the growing potential and value for hydrogels in tissue engineering. The fabrication of hydrogel-based cellular scaffolds for the generation of bioengineered tissues has been based on knowledge of the composition and structure of ECM. Attempts at recreating ECM have used either naturally-derived ECM components or synthetic polymers with structural integrity derived from hydrogels. Due to their increasing use, their biocompatibility has been questioned since the use of these biomaterials needs to be effective and safe. It is not surprising then that the evaluation of biocompatibility of these types of biomaterials for regenerative and tissue engineering applications has been expanded from being primarily investigated in a laboratory setting to being applied in the multi-billion dollar medicinal industry. This review will aid in the improvement of design of non-invasive, smart hydrogels that can be utilized for tissue engineering and other biomedical applications. In this review, the biocompatibility of hydrogels and design criteria for fabricating effective scaffolds are examined. Examples of natural and synthetic hydrogels, their biocompatibility and use in tissue engineering are discussed. The merits and clinical complications of hydrogel scaffold use are also reviewed. The article concludes with a future outlook of the field of biocompatibility within the context of hydrogel-based scaffolds.

Oral, ultra-long-lasting drug delivery: Application toward malaria elimination goals.

Efforts at elimination of scourges, such as malaria, are limited by the logistic challenges of reaching large rural populations and ensuring patient adherence to adequate pharmacologic treatment. We have developed an oral, ultra-long-acting capsule that dissolves in the stomach and deploys a star-shaped dosage form that releases drug while assuming a geometry that prevents passage through the pylorus yet allows passage of food, enabling prolonged gastric residence. This gastric-resident, drug delivery dosage form releases small-molecule drugs for days to weeks and potentially longer. Upon dissolution of the macrostructure, the components can safely pass through the gastrointestinal tract. Clinical, radiographic, and endoscopic evaluation of a swine large-animal model that received these dosage forms showed no evidence of gastrointestinal obstruction or mucosal injury. We generated long-acting formulations for controlled release of ivermectin, a drug that targets malaria-transmitting mosquitoes, in the gastric environment and incorporated these into our dosage form, which then delivered a sustained therapeutic dose of ivermectin for up to 14 days in our swine model. Further, by using mathematical models of malaria transmission that incorporate the lethal effect of ivermectin against malaria-transmitting mosquitoes, we demonstrated that this system will boost the efficacy of mass drug administration toward malaria elimination goals. Encapsulated, gastric-resident dosage forms for ultra-long-acting drug delivery have the potential to revolutionize treatment options for malaria and other diseases that affect large populations around the globe for which treatment adherence is essential for efficacy.

Physicochemical characterization of a navy bean (Phaseolus vulgaris) protein fraction produced using a solvent-free method.

A solvent-free electrostatic separation method was employed to separate navy bean flour (NBF) into protein-rich (PR) and starch-rich (SR) fractions. The physicochemical properties of NBF and separated fractions were compared to proteins (navy bean isolate (NBI) and 7S globulin) prepared using a wet process. Gel electrophoresis confirmed that the protein distribution in the isolated fractions was similar to that of NBF. The protein profile of NBI and 7S globulin was found to be devoid of certain proteins that were found in the NBF and PR fraction. Amino acid analysis revealed that the NBI and 7S globulin had a lower content of sulfur-containing amino acids compared to NBF and the electrostatically isolated fractions. CD and fluorescence spectroscopy confirmed that denaturation of the proteins during the acid precipitation is likely. This novel solvent-free electrostatic separation process preserves the native protein structure found in NBF and improves the recovery of some of the smaller MW proteins.

DEGylation Enhanced the Stability of Peptide-siRNA Complexes in Serum.

Small interfering RNA (siRNA) shows great therapeutic potential due to its ability to regulate gene expression in a highly selective manner, but this application has been limited by effective delivery, partly because of the low nuclease resistance of siRNA in the presence of serum, and inefficient cellular uptake. We previously reported a library of cell-penetrating and amino acid-pairing peptides that facilitate effective siRNA delivery to mammalian cells without causing cytotoxicity, but they are unstable within serum-containing medium. Here, we investigated the possibility of conjugating the peptide with diethylene glycol to improve its serum stability without compromising its gene-regulation capability. One of the most promising peptides, C6M1, was conjugated with diethylene glycol, and its incorporated siRNA complexes had excellent serum stability and highly efficient cellular uptake with negligible cytotoxicity. The gene-silencing ability of diethylene glycol conjugated-peptide/siRNA complexes was comparable to that of non-conjugated peptide/siRNA at both mRNA and protein levels. Our data demonstrate that conjugating peptides with diethylene glycol is a promising method for improving siRNA delivery by improving its serum stability.

Natural Carriers for siRNA Delivery.

This review is based on carriers of natural origin such as polysaccharides, proteins, and cell derived entities which have been used for delivery of siRNA. To realize the therapeutic potential of a delivery system, the role of the carrier is of utmost importance. Historical aspects of viral vectors, the first carriers of genes are briefly outlined. Chitosan, one of the extensively experimented carriers, alginates and other polysaccharides have shown success in siRNA delivery. Peptides of natural origin and mimics thereof have emerged as another versatile carrier. Exosomes and mini cells of cellular origin are the newest entrants to the area of siRNA delivery and probably the closest one can get to a natural carrier. In many of the carriers, modifications have provided better efficiency in delivery. The salient features of the carriers and their advantages and disadvantages are also reviewed.

A novel peptide for efficient siRNA delivery in vitro and therapeutics in vivo.

Small interfering RNA (siRNA) shows great therapeutic potential due to its ability to regulate gene expression in a highly selective manner. However, its application has been limited by ineffective cellular uptake of siRNAs. To achieve successful gene-silencing efficiency, a safe and effective delivery vector is generally required. In this study, we designed a series of amphipathic peptides that comprised a variant of a nuclear localization sequence, 0-6 histidine residues and an optional stearic acid group. Among these candidates, STR-HK exhibited good characteristics as a safe and efficient siRNA delivery vector, facilitating efficient siRNA delivery to mammalian cells without causing cytotoxicity. Moreover, the intratumoral injection of STR-HK/siRNA complexes achieved high anti-tumor activity through the downregulation of the Bcl-2 protein in mice, with an inhibition rate of 62.8%. Our data demonstrate that STR-HK is a highly promising siRNA delivery vector for therapeutic applications.

Evaluation of biocompatibility of the AC8 peptide and its potential use as a drug carrier.

Peptide-based nanoparticles have emerged as promising drug delivery systems for targeted cancer therapy. Yet, the biocompatibility of these nanoparticles has not been elucidated. Here, the in vitro biocompatibility and toxicity and in vivo immunocompatibility and bioactivity of the self/coassembling peptide AC8 in its nanoparticle form are evaluated. AC8 showed minimal hemolytic activity (5%) and did not cause aggregation of red blood cells. The in vitro assay revealed that AC8 did not activate the complement system via the classical or alternative pathway but did activate the lectin pathway to a small extent. However, AC8 showed no C3a and C5a anaphylotoxin activation suggesting that complement activation did not proceed to the later, inflammatory, stages. The in vivo immune response assay showed that administration of AC8 to BALB/c mice had no effect on the weight of immune organs or body weight of mice at doses less than 0.1 mg/kg. This peptide also did not have any effect on the expression of CD3+ T-cells and natural killer (NK) cells, the ratio of CD4+/CD8+ T-cell, and the proliferation of B-cells. These results suggest that AC8 can be a potential carrier candidate for drug delivery.

Serum stability and physicochemical characterization of a novel amphipathic peptide C6M1 for siRNA delivery.

The efficient delivery of nucleic acids as therapeutic agents is a major challenge in gene therapy. Peptides have recently emerged as a novel carrier for delivery of drugs and genes. C6M1 is a designed amphipathic peptide with the ability to form stable complexes with short interfering RNA (siRNA). The peptide showed a combination of random coil and helical structure in water but mainly adopted a helical conformation in the presence of anions or siRNA. Revealed by dynamic light scattering (DLS) and microscopy techniques, the interaction of C6M1 and siRNA in water and HEPES led to complexes of ∼70 and ∼155 nm in size, respectively, but showed aggregates as large as ∼500 nm in PBS. The time-dependent aggregation of the complex in PBS was studied by DLS and fluorescence spectroscopy. At molar ratio of 15∶1, C6M1 was able to completely encapsulate siRNA; however, higher molar ratios were required to obtain stable complexes. Naked siRNA was completely degraded in 4 h in the solution of 50% serum; however C6M1 protected siRNA against serum RNase over the period of 24 h. Western blotting experiment showed ∼72% decrease in GAPDH protein level of the cells treated with C6M1-siRNA complexes while no significant knockdown was observed for the cells treated with naked siRNA.

In vitro and in vivo therapeutic siRNA delivery induced by a tryptophan-rich endosomolytic peptide.

At the forefront of medicine, gene therapy provides an effective way to treat a range of diseases by regulating defective genes at the root of the disease. Short interfering RNAs (siRNAs) hold great promise as therapeutic agents in this domain; however, intracellular delivery remains a major obstacle to clinical applications of therapeutic siRNAs. Here we report a peptide designed to mediate siRNA delivery. This peptide, C6M1, is rationally designed to promote the endosomal escape ability of an existing peptide sequence. Formed C6M1-siRNA nanoscale complexes are able to deliver siRNA into cells and induce specific gene knockdown with low toxicity. The increased membrane disruption ability under acidic conditions of the peptide with tryptophan residue substitution may contribute to the enhanced gene silence efficacy. Intratumoral injection of the complexes results in a marked reduction of tumor growth through downregulation of antiapoptotic Bcl-2 protein in mice. In addition, the C6M1-siRNA complex was proven safe at transfection concentration by cytotoxicity assay. These results demonstrate that the C6M1-siRNA complex is a potent system for efficient gene delivery in vitro and in vivo.

Modification of a designed amphipathic cell-penetrating peptide and its effect on solubility, secondary structure, and uptake efficiency.

The development of safe and efficient nonviral gene delivery carriers has received a great deal of attention in the past decade. A class of amphipathic peptides has shown to be able to cross cell membranes and deliver cargo to the intracellular environment. Here, we introduce an 18-mer amphipathic peptide, C6M1, as a modified version of peptide C6 for short interfering RNA (siRNA) delivery. The importance of tryptophan residues and the effect of peptide sequence modification on its solubility, secondary structure, cytotoxicity, and uptake efficiency were investigated. The solubility of C6M1 in aqueous solutions was greatly enhanced compared to that of C6, confirmed by surface tension and anilinonaphthalene-8-sulfonic acid fluorescence measurements. C6M1 had a random/helical structure in water with the ability to attain a helical conformation in the presence of anionic components or membrane-mimicking environments. The modification significantly reduced the cytotoxicity of the peptide, making it a safer carrier for siRNA delivery. C6M1 was also found ∼90% more efficient than C6 in delivering Cy3-labeled siRNA in Chinese hamster ovary cells.

Biocompatibility of engineered nanoparticles for drug delivery.

The rapid advancement of nanotechnology has raised the possibility of using engineered nanoparticles that interact within biological environments for treatment of diseases. Nanoparticles interacting with cells and the extracellular environment can trigger a sequence of biological effects. These effects largely depend on the dynamic physicochemical characteristics of nanoparticles, which determine the biocompatibility and efficacy of the intended outcomes. Understanding the mechanisms behind these different outcomes will allow prediction of the relationship between nanostructures and their interactions with the biological milieu. At present, almost no standard biocompatibility evaluation criteria have been established, in particular for nanoparticles used in drug delivery systems. Therefore, an appropriate safety guideline of nanoparticles on human health with assessable endpoints is needed. In this review, we discuss the data existing in the literature regarding biocompatibility of nanoparticles for drug delivery applications. We also review the various types of nanoparticles used in drug delivery systems while addressing new challenges and research directions. Presenting the aforementioned information will aid in getting one step closer to formulating compatibility criteria for biological systems under exposure to different nanoparticles.

A new amphipathic, amino-acid-pairing (AAP) peptide as siRNA delivery carrier: physicochemical characterization and in vitro uptake.

RNA interference has emerged as a powerful tool in biological and pharmaceutical research; however, the enzymatic degradation and polyanionic nature of short interfering RNAs (siRNAs) lead to their poor cellular uptake and eventual biological effects. Among nonviral delivery systems, cell-penetrating peptides have been recently employed to improve the siRNA delivery efficiency. Here we introduce an 18-mer amphipathic, amino-acid-pairing peptide, C6, as an siRNA delivery carrier. Peptide C6 adopted a helical structure upon coassembling with siRNA. The C6-siRNA coassembly showed a size distribution between 50 and 250 nm, confirmed by dynamic light scattering and atomic force microscopy. The C6-siRNA interaction enthalpy and stoichiometry were 8.8 kJ·mol(-1) and 6.5, respectively, obtained by isothermal titration calorimetry. A minimum C6/siRNA molar ratio of 10:1 was required to form stable coassemblies/complexes, indicated by agarose gel shift assay and fluorescence spectroscopy. Peptide C6 showed lower toxicity and higher efficiency in cellular uptake of siRNA compared with Lipofectamine 2000. Fluorescence microscopy images also confirmed the localization of C6-siRNA complexes in the cytoplasm using Cy3-labeled siRNAs. These results indicate high capabilities of C6 in forming safe and stable complexes with siRNA and enhancing its cellular uptake.

Self-assembling peptides: potential role in tumor targeting.

This review focuses on the application of two classes of peptides, i.e., self-assembling peptides (SAPs) and cell-targeting peptides (CTPs), in the development of nanocarrier delivery systems. Self-assembling peptides are emerging in a wide range of biomedical and bioengineering applications and fall into several classes, including peptide amphiphilies, bolaamphiphile peptides, cyclic peptides, and ionic complementary peptides, which can be found naturally or synthesized. The advantage of synthesizing peptides is that their self-assembling properties can be exploited to form desirable structures for various applications. Another, unique property of self-assembling peptides, is stimuli-responsibility in different environments including various pHs, temperatures, ionic strengths, etc. These characteristics make peptides applicable in a wide range of biomaterials in drug discovery. This study reviews the design principles of well-known self-assembling peptides, as well as their physical/chemical properties. In addition, it discusses the therapeutic cancer-targeting peptides and current combinatorial peptide library methods used to identify targeting peptides. Cancer-targeting peptides can target either tumor cell surfaces or tumor vasculature. The RGD peptide is one of the first tumor-targeting peptides that can bind to self-assembling peptides or any other nanocarrier to improve the therapeutic efficiency of targeting drug delivery systems.

Peptide mediated siRNA delivery.

Applying RNA interference to silence a specific gene has opened a new and promising avenue of gene therapy. But a key bottleneck is the poor stability and inability of naked siRNA to translocate through cell membranes. Among several delivery systems, cationic peptides capable of penetrating cell membranes have drawn attention due to their structural and functional versatility, potential biocompatibility and ability to target cells. In this review, different classes of peptides employed in siRNA delivery are reviewed. In particular, a new class of siRNA delivery peptides with high transfection efficiency and low cytotoxicity is introduced.

Physicochemical characterization of siRNA-peptide complexes.

Short interfering RNAs (siRNAs) trigger RNA interference (RNAi), where the complementary mRNA is degraded, resulting in silencing of the encoded protein. A delivery carrier is desired to increase the solution stability of siRNA and improve its cellular uptake to overcome its rapid enzymatic degradation and low transfection efficiency. In this study, Arginine-9 (R9), a cell-penetrating peptide derived from the HIV 1 Tat protein, was investigated as a potential carrier for siRNAs. A connective tissue growth factor (CTGF) encoding siRNA was used because of its therapeutic potential of treating breast cancer. The interaction between R9 and siRNA was studied by UV/vis spectroscopy and circular dichroism (CD). The hydrodynamic diameter of the siRNA-R9 complexes was determined by dynamic light scattering (DLS), and the Zeta potential of the complexes was obtained by measuring the electrophoretic mobility. The effect of salt addition is also quantified using UV-vis spectroscopy. The siRNA and R9 readily formed complexes/aggregates through molecular association, accompanying a change in surface charge with increasing peptide concentration, reaching a maximum hydrodynamic diameter of approximately 1 mum at siRNA saturation. The highest binding ratio of R9 to siRNA determined from the UV/vis spectra and CD is 10.3:1 and 39.1:1 from DLS (corresponds to charge ratios of 2.2:1 (+/-) and 8.4:1, respectively). The difference in binding ratio is possibly because of the difference in signal contribution between absorption and light scattering. The physicochemical characterization of CTGF siRNA-R9 complexes presented here have shown that various methods can be used to control the properties of the siRNA-peptide complexes, which provide a basis for the formulation of siRNA therapeutics with peptide carriers.