Multiple Bioactive Agents Research Articles

Zinc and chitosan-enhanced β-tricalcium phosphate from calcined fetal bovine bone for mandible reconstruction.

Mandibular defects pose significant challenges in reconstructive surgery, and scaffold materials are increasingly recognized for their potential to address these challenges. Among various scaffold materials, Beta-tricalcium phosphate (β-TCP) is noted for its exceptional osteogenic properties. However, improvements in its biodegradation rate and mechanical strength are essential for optimal performance. In this study, we developed a novel β-TCP-based scaffold, CFBB, by calcining fetal bovine cancellous bone. To enhance its properties, we modified CFBB with Chitosan (CS) and Zinc (Zn), creating three additional scaffold materials: CFBB/CS, CFBB/Zn2+, and CFBB/Zn2+/CS. We conducted comprehensive assessments of their physicochemical and morphological properties, degradation rates, biocompatibility, osteogenic ability, new bone formation, and neovascularization both in vitro and in vivo. Our findings revealed that all four materials were biocompatible and safe for use. The modifications with CS and Zn2+ significantly improved the mechanical strength, osteogenic, and angiogenic properties of CFBB, while concurrently decelerating its resorption rate. Among the tested materials, CFBB/Zn2+/CS demonstrated superior performance in promoting bone regeneration and vascularization, making it a particularly promising candidate for mandibular reconstruction. The CFBB/Zn2+/CS scaffold material, with its enhanced mechanical, osteogenic, and angiogenic properties, and a controlled resorption rate, emerges as a highly effective alternative for the repair of oral mandible defects. This study underscores the potential of combining multiple bioactive agents in scaffold materials to improve their functionality for specific clinical applications in bone tissue engineering.

Pro-vasculogenic Fibers by PDA-Mediated Surface Functionalization Using Cell-Free Fat Extract (CEFFE).

Formation of adequate vascular network within engineered three-dimensional (3D) tissue substitutes postimplantation remains a major challenge for the success of biomaterials-based tissue regeneration. To better mimic the in vivo angiogenic and vasculogenic processes, nowadays increasing attention is given to the strategy of functionalizing biomaterial scaffolds with multiple bioactive agents. Aimed at engineering electrospun biomimicking fibers with pro-vasculogenic capability, this study was proposed to functionalize electrospun fibers of polycaprolactone/gelatin (PCL/GT) by cell-free fat extract (CEFFE or FE), a newly emerging natural "cocktail" of cytokines and growth factors extracted from human adipose tissue. This was achieved by having the electrospun PCL/GT fiber surface coated with polydopamine (PDA) followed by PDA-mediated immobilization of FE to generate the pro-vasculogenic fibers of FE-PDA@PCL/GT. It was found that the PDA-coated fibrous mat of PCL/GT exhibited a high FE-loading efficiency (∼90%) and enabled the FE to be released in a highly sustained manner. The engineered FE-PDA@PCL/GT fibers possess improved cytocompatibility, as evidenced by the enhanced cellular proliferation, migration, and RNA and protein expressions (e.g., CD31, vWF, VE-cadherin) in the human umbilical vein endothelial cells (huvECs) used. Most importantly, the FE-PDA@PCL/GT fibrous scaffolds were found to enormously stimulate tube formation in vitro, microvascular development in the in ovo chick chorioallantoic membrane (CAM) assay, and vascularization of 3D construct in a rat subcutaneous embedding model. This study highlights the potential of currently engineered pro-vasculogenic fibers as a versatile platform for engineering vascularized biomaterial constructs for functional tissue regeneration.

Preparation and Characterization of Sodium Alginate-reduced Graphene Oxide Microbeads for Co-Delivery of Doxorubicin and 5-Fluorouracil

Combination therapy has gained significant interest in recent years due to its potential to produce greater therapeutic success than mono-drug therapy. However, carriers capable of effective and stimuli-responsive codelivery of several drugs are severely lacking at the time. To address this need, the study represents the fabrication of reduced graphene oxide-sodium alginate microbeads for co-delivery and controlled release of doxorubicin (DOX) and 5-fluorouracil (5-FU). The developed beads were characterized by Fourier transform infrared spectroscopy (FTIR), Differential scanning calorimetry (DSC), Thermogravimetric analysis (TGA), X-ray diffraction (XRD) and Scanning electron microscopy (SEM). Swelling and in vitro drug release experiments were performed at pH 7.4, 5.4 and 1.2 at 37 °C. The findings indicated that the pH of the test media could be appropriate for intestinal drug delivery. The release process was studied with the release data being integrated into the Korsmeyer-Peppas equation. The presence of rGO in the polymeric matrix influences the drug release rate. The anticancer activity of the alginate beads was assessed using an MTT assay against MCF-7 cells. The findings indicate that the dual-drug containing microbeads showed good activity compared to 5-FU loaded microbeads. The results showed that the developed microbeads warrant further development for applications that require simultaneous administration of multiple bioactive agents.

Preparation and characterization of 2-hydroxyethyl starch microparticles for co-delivery of multiple bioactive agents

The present study reports the generation of 2-hydroxyethyl starch microparticles for co-delivery and controlled release of multiple agents. The obtained microparticles are characterized by using Fourier transform infrared spectroscopy, differential scanning calorimetry, X-ray diffraction analysis, energy-dispersive X-ray spectroscopy, and scanning electron microscopy. By using ofloxacin and ketoprofen as drug models, the release sustainability of the microparticles is examined at pH 1.2, 5.4, and 6.8 at 37 °C, with Fickian diffusion being found to be the major mechanism controlling the kinetics of drug release. Upon being loaded with the drug models, the microparticles show high efficiency in acting against Escherichia coli and Bacillus cereus. The results suggest that our reported microparticles warrant further development for applications in which co-administration of multiple bioactive agents is required.

Interfacial Stacks of Polymeric Nanofilms on Soft Biological Surfaces that Release Multiple Agents.

We report a general and facile method that permits the transfer (stacking) of multiple independently fabricated and nanoscopically thin polymeric films, each containing a distinct bioactive agent, onto soft biomedically relevant surfaces (e.g., collagen-based wound dressings). By using polyelectrolyte multilayer films (PEMs) formed from poly(allyl amine hydrochloride) and poly(acrylic acid) as representative polymeric nanofilms and micrometer-thick water-soluble poly(vinyl alcohol) sacrificial films to stack the PEMs, we demonstrate that it is possible to create stacked polymeric constructs containing multiple bioactive agents (e.g., antimicrobial and antibiofilm agents) on soft and chemically complex surfaces onto which PEMs cannot be routinely transferred by stamping. We illustrate the characteristics and merits of the approach by fabricating stacks of Ga3+ (antibiofilm agent)- and Ag+ (antimicrobial agent)-loaded PEMs as prototypical examples of agent-containing PEMs and demonstrate that the stacked PEMs incorporate precise loadings of the agents and provide flexibility in terms of tuning release rates. Specifically, we show that simultaneous release of Ga3+ and Ag+ from the stacked PEMs on collagen-based wound dressings can lead to synergistic effects on bacteria, killing and dispersing biofilms formed by Pseudomonas aeruginosa (two strains: ATCC 27853 and MPAO1) at sufficiently low loadings of agents such that cytotoxic effects on mammalian cells are avoided. The approach is general (a wide range of bioactive agents other than Ga3+ and Ag+ can be incorporated into PEMs), and the modular nature of the approach potentially allows end-user functionalization of soft biological surfaces for programmed release of multiple bioactive agents.

Co-delivery of doxorubicin and 131I by thermosensitive micellar-hydrogel for enhanced in situ synergetic chemoradiotherapy

Combined chemoradiotherapy is potent to defeat malignant tumor. Concurrent delivery of radioisotope with chemotherapeutic drugs, which also act as the radiosensitizer, to tumor tissues by a single vehicle is essential to achieve this objective. To this end, a macroscale injectable and thermosensitive micellar-hydrogel (MHg) depot was constructed by thermo-induced self-aggregation of poly(ε-caprolactone-co-1,4,8-trioxa[4.6]spiro-9-undecanone)-poly(ethyleneglycol)-poly(ε-caprolactone-co-1,4,8-trioxa[4.6]spiro-9-undecanone) (PECT) triblock copolymer micelles (Ms), which could not only serve as a micellar drug reservoir to locally deliver concentrated nano chemotherapeutic drugs, but also immobilize radioisotopes at the internal irradiation hot focus. Doxorubicin (DOX) and iodine-131 labeled hyaluronic acid (131I-HA) were used as the model therapeutic agents. The aqueous mixture of drug-loaded PECT micelles and 131I-HA exhibited sol-to-gel transition around body temperature. In vitro drug release study indicated that PECT/DOX Ms were sustainedly shed from the native PECT/DOX MHg formulation, which could be internalized by tumor cells with rapid intracellular DOX release. This hydrogel formulation demonstrated considerable in vitro antitumor effect as well as remarkable radiosensitization. In vivo subcutaneous injection of PECT MHg demonstrated that 131I isotope was immobilized stably at the injection location and no obvious indication of damage to major organs were observed as indicated by the histopathological analysis. Furthermore, the peritumoral injection of chemo-radiation therapeutic agents-encapsulated MHg formulation on tumor-bearing nude mice resulted in the desired combined treatment effect, which significantly improved the tumor growth inhibition efficiency with minimized drug-associated side effects to major organs. Consequently, such a thermosensitive MHg formulation, which enabled the precise control over the dosage and ratio of combination therapeutic agents to obtain the desired therapeutic effect with a single drug administration and reduced side effects, holds great potential for spatiotemporally delivery of multiple bioactive agents for sustained combination therapy.

Inhibitory activity of extracts of Hebridean brown seaweeds on lipase activity.

The effect of three Hebridean brown seaweeds on lipase activity was assessed using a turbidimetric lipase activity assay and an in vitro simulation of the upper digestive tract. The preparations of Ascophyllum nodosum, Fucus vesiculosus, and Pelvetia canaliculata were tested; whole seaweed homogenate, sodium carbonate extract, and ethanol extracts (pellet and supernatant were tested separately). All extracts showed significant inhibition of lipase, suggesting multiple bioactive agents, potentially including alginates, fucoidans, and polyphenols. Whole homogenate extract of F. vesiculosus was the most potent inhibitor of Lipase (IC50 = 0.119 mg mL-1), followed by ethanol supernatant (IC50 = 0.159 mg mL-1) while ethanol pellet and sodium carbonate extract showed relatively weaker inhibition (IC50 = 0.360 mg mL-1 and IC50 = 0.969 mg mL-1 respectively). For A. nodosum and P. canaliculata, strongest inhibition occurred with ethanol pellet (IC50 = 0.238 and 0.228 mg mL−1, respectively). These inhibitory effects were validated in a model gut system. The data presented herein suggests the use of seaweed as a potential weight management tool is deserving of further investigation.

An injectable, thermosensitive and multicompartment hydrogel for simultaneous encapsulation and independent release of a drug cocktail as an effective combination therapy platform

Combination chemotherapy is potent to combat diseases. Simultaneous and segregated delivery of multiple drugs in a single vehicle is essential to achieve this objective. In the present study, an injectable, thermosensitive and multicompartment hydrogel (MCH) was developed by the facile cooperative and incompatible assembly of PEGylated hydrocarbon nanoparticles with PEGylated fluorocarbon nanoparticles. The cooperative assembly behavior was investigated by fluorescence resonance energy transfer (FRET) technology, and the result demonstrated that the incompatible nanoparticle cores possibly accounted for the multicompartment formation in hydrogel. Paclitaxel and doxorubicin could be easily and separately integrated into the different compartments of MCH serving as a sustained drug cocktail formulation. In vitro drug release indicated drugs were liberated in a simultaneous but independent manner without any effect on each other. In vitro and in vivo antitumor activity indicated that peritumoral injection of drug cocktail encapsulated MCH formulation could well achieve the combination effect, which significantly improved the tumor growth inhibition efficiency as well as minimized the drug-associated side effects compared to intravenous injection of free drug cocktail. Furthermore, such a delivery device would allow precise adjustment of drug dosage to the desired effect, achieve spatial–temporal simultaneous and synchronized presence of combination drugs in the target tissue and obviate repeated drug administrations to improve patient compliance. The thermosensitive multicompartment hydrogel cocktail formulation holds great promise for simultaneous and segregated delivery of multiple bioactive agents for sustained combination therapy.

PLGA/gelatin hybrid nanofibrous scaffolds encapsulating EGF for skin regeneration.

The novel strategies of skin regenerative treatment are aimed at the development of biologically responsive scaffolds capable of delivering multiple bioactive agents and cells to the target tissues. In this study, nanofibers of poly(lactic-co-glycolic acid) (PLGA) and gelatin were electrospun and the effect of parameters viz polymer concentration, acid concentration, flow rate and voltage on the morphology of the fibers were investigated. PLGA nanofibers encapsulating epidermal growth factor were also prepared through emulsion electrospinning. The core-sheath structure of the nanofibers was verified by transmission electron microscopy. The hemostatic attributes and the biocompatibility of the scaffolds for human fibroblast cell were scrutinized. Furthermore, gene expression of collagen type I and type III by the cells on the scaffolds was quantified using real-time reverse transcriptase polymerase chain reaction. The results indicated desirable bioactivity and hemostasis of the scaffolds with the capability of encapsulation and controlled release of the protein which can be served as skin tissue engineering scaffolds and wound dressings.

In-situ crosslinking hydrogels for combinatorial delivery of chemokines and siRNA–DNA carrying microparticles to dendritic cells

Polymer-based, injectable systems that can simultaneously deliver multiple bioactive agents in a controlled manner could significantly enhance the efficacy of next generation therapeutics. For immunotherapies to be effective, both prophylactically or therapeutically, it is not only critical to drive the antigen (Ag)-specific immune response strongly towards either T helper type 1 (Th1) or Th2 phenotype, but also to promote recruitment of a high number of antigen-presenting cells (APCs) at the site of immunization. We have recently reported a microparticle-based system capable of simultaneously delivering siRNA and DNA to APCs. Here we present an in-situ crosslinkable, injectable formulation containing dendritic cell (DC)-chemo-attractants and dual-mode DNA–siRNA loaded microparticles to attract immature DCs and simultaneously deliver, to the migrated cells, immunomodulatory siRNA and plasmid DNA antigens. These low crosslink density hydrogels were designed to degrade within 2–7 days in vitro and released chemokines in a sustained manner. Chemokine carrying gels attracted 4–6 folds more DCs over a sustained period in vitro, compared to an equivalent bolus dose. Interestingly, migrated DCs were able to infiltrate the hydrogels and efficiently phagocytose the siRNA–DNA carrying microparticles. Hydrogel embedded microparticles co-delivering Interleukin-10 siRNA and plasmid DNA antigens exhibited efficient Interleukin-10 gene knockdown in migrated primary DCs in vitro.

Use of Degradable and Nondegradable Nanomaterials for Controlled Release

Drug-delivery devices are fundamentally important in improving the pharmacological profiles of therapeutic molecules. Nanocontrolled-release systems are attracting a lot of attention currently owing to their large surface area and their ability to target delivery to specific sites in the human body. In addition, they can penetrate the cell membrane for gene, nucleic acid and bioactive peptide/protein delivery. Representative applications of nanodrug-delivery systems include controlled-release wound dressings, controlled-release scaffolds for tissue regeneration and implantable biodegradable nanomaterial-based medical devices integrated with drug-delivery functions. We review the present status and future perspectives of various types of nanocontrolled-release systems. Although many of the well-established degradable and nondegradable controlled-release vehicles are being investigated for their processing into nanocarriers, several new emerging nanomaterials are being studied for their controlled-release properties. The release of multiple bioactive agents, each with its own kinetic profile, is becoming possible. In addition, integration of the nanocontrolled-release systems with other desirable functions to create new, cross-discipline applications can also be realized.

In‐vitro assays of polymer‐coated stents eluting platelet glycoprotein IIb/IIIa receptor monoclonal antibody

The monoclonal antibody (mAb) to the platelet glycoprotein (GP) IIb/IIIa receptor has potent antiplatelet and antithrombotic characteristics shown to reduce thrombus-related major complications after coronary angioplasty. This mAb can be incorporated in drug-eluting stents capable of releasing single or multiple bioactive agents into the bloodstream and surrounding tissues. Stents eluting the monoclonal mouse anti-human platelet glycoprotein IIb/IIIa antibody SZ-262 were tested for their effectiveness in improving the blood compatibility and the antithrombotic characteristics by immunofluorescence and scanning electron microscopy (SEM). The SEM results convincingly demonstrated that the surface of the mAb eluting-stents was completely free of platelet uptake without any sign of cellular debris or proteinaceous deposits, compared with controls. The deformation index of platelets on the L-polylactic acid (L-PLA) coated stents were higher than bare Nitinol intravascular stents, as shown by SEM images. Monoclonal antibody to the platelet GP IIb/IIIa receptor, when eluting from L-PLA polymer-coated stents, effectively inhibits platelet aggregation in the stent microenvironment, thus demonstrating a potential capacity of reducing thrombosis, improving blood flow and arterial patency rates, and inhibiting cyclic blood flow variations. These results highlight the possibility of such monoclonal antibody-eluting stents to reduce or possibly eliminate thrombosis and in-stent restenosis.

New Drug-Eluting Stents

The introduction of drug-eluting stents (DES) has rapidly and profoundly affected the field of interventional cardiology, and DESs are now used in a majority of intracoronary stenting procedures. As a result of the innumerable “trial-and-error” endeavors, DESs have emerged as a potential solution for solving the problem of restenosis. DESs are coated stents capable of releasing single or multiple bioactive agents into the bloodstream and the surrounding tissues. The efficacy and safety of DESs might differ depending on the pharmacologic agents and stent delivery systems that are used. Recent research has focused on the various constituents of DESs, including the stent backbone, the materials used as drug-delivery vehicles, and the physicochemical properties of the pharmacotherapeutic agents themselves. Stent-induced mechanical arterial injury and a foreign body response to the prosthesis incite acute and chronic inflammation in the vessel wall, along with the elaboration of cytokines and growth factors that induce multiple signaling pathways to activate smooth muscle cell migration and proliferation. Utilization of antiproliferative agents delivered locally via the drug-eluting stents has dramatically reduced the stent restenosis rate. The polymer-regulated delivery of both paclitaxel and sirolimus at the site of arterial injury has been shown to reduce the clinical and angiographic restenosis rates after stent implantation in de novo coronary lesion. Other DESs have yielded somewhat less brilliant results or even true failures, while a number of new drugs and new stent platforms are now under clinical or preclinical evaluation. In this review, we describe the main clinical trials on DESs and the most recent information that has been derived from observational studies and registries. Moreover, the preliminary results on the new DESs are also summarized. (Korean Circulation J 2005;35:197-205)

Synthesis of a prodrug designed to release multiple inhibitors of the epidermal growth factor receptor tyrosine kinase and an alkylating agent: a novel tumor targeting concept.

The synthesis of a novel acetoxymethyltriazene designed to be a prodrug of multiple inhibitors of the epidermal growth factor receptor (EGFR) and a methyldiazonium species is described. Studies with each of the expected metabolites demonstrated significant EGFR tyrosine kinase inhibitory activities and the released methyldiazonium was trapped with p-nitrobenzylpyridine. Their ability to damage genomic DNA in whole cells was demonstrated by using the single cell microelectrophoresis (comet) assay. The results suggest that this approach may well represent a novel drug combination strategy involving single molecules masking multiple bioactive agents.

New Frontiers in Cardiology

Stents represent a major advance in the treatment of obstructive coronary artery disease since the advent of balloon angioplasty. The number of percutaneous coronary interventions performed each year has expanded considerably since the early days. Angioplasty procedures doubled in Europe between 1992 and 1996,1 while an estimated 601 000 percutaneous coronary revascularizations were performed in the United States in 1997.2 Unfortunately, many of these patients develop an exaggerated vascular neointimal proliferation after stenting—namely, in-stent restenosis. Much research has been devoted to the pathophysiology and treatment of in-stent restenosis. As a result of many relentless “trial-and-error” endeavors, drug-eluting stents have emerged as a potential solution for restenosis. Drug-eluting stents are coated stents capable of releasing single or multiple bioactive agents into the bloodstream and surrounding tissues. We have been commissioned to present an overview on drug-eluting stents. Acknowledging the challenge of examining such a dynamic and flourishing field, our goals in this 2-part article were to provide a broad perspective of the development of drug-eluting stent technology, to summarize the available clinical data, and to introduce emerging concepts for the understanding and application of this new device in clinical practice. In 1991, stent use was still facing skepticism because of an unacceptably high (20% to 25%) incidence of thrombotic complications.3 Systemic anticoagulation proved disappointing in reducing the catastrophic consequences of stent thrombosis, such as myocardial infarction and sudden death. Consequently, antithrombotic stent coatings were developed to decrease the inherent thrombogenicity of coronary metallic stents. Some heparin-coated stents have become available for clinical use: BX-Velocity Carmeda-coated stent (Johnson & Johnson), Wiktor Hepamed-coated stent (Medtronic, Inc), and the Jostent Corline-coated stent (Jomed International AB). Heparin-coated stents differ from drug-eluting stents because the medication is covalently bonded to the device and hence may remain attached long after deployment. However, these stents …