Bone-specific Drug Delivery Research Articles

Bone-Targeted Nanoparticle Drug Delivery System-Mediated Macrophage Modulation for Enhanced Fracture Healing.

Despite decades of progress, developing minimally invasive bone-specific drug delivery systems (DDS) to improve fracture healing remains a significant clinical challenge. To address this critical therapeutic need, nanoparticle (NP) DDS comprised of poly(styrene-alt-maleic anhydride)-b-poly(styrene) (PSMA-b-PS) functionalized with a peptide that targets tartrate-resistant acid phosphatase (TRAP) and achieves preferential fracture accumulation has been developed. The delivery of AR28, a glycogen synthase kinase-3 beta (GSK3β) inhibitor, via the TRAP binding peptide-NP (TBP-NP) expedites fracture healing. Interestingly, however, NPs are predominantly taken up by fracture-associated macrophages rather than cells typically associated with fracture healing. Therefore, the underlying mechanism of healing via TBP-NP is comprehensively investigated herein. TBP-NPAR28 promotes M2 macrophage polarization and enhances osteogenesis in preosteoblast-macrophage co-cultures in vitro. Longitudinal analysis of TBP-NPAR28 -mediated fracture healing reveals distinct spatial distributions of M2 macrophages, an increased M2/M1 ratio, and upregulation of anti-inflammatory and downregulated pro-inflammatory genes compared to controls. This work demonstrates the underlying therapeutic mechanism of bone-targeted NP DDS, which leverages macrophages as druggable targets and modulates M2 macrophage polarization to enhance fracture healing, highlighting the therapeutic benefit of this approach for fractures and bone-associated diseases.

Multifunctional bone substitute using carbon dot and 3D printed calcium-deficient hydroxyapatite scaffolds for osteoclast inhibition and fluorescence imaging.

Multifunctional bone substitute materials (BSM) have gained considerable attention with the exponential increase in aging populations. The development of hybrid materials for diagnosis and therapy of bone-related diseases and dysfunctions, especially, has been a significant challenge in the biological and the biomedical field, due to the shortage of agents with specificity and selectivity toward bone. In this study, a hybrid material, referred as Alen-CDs@CDHA, fabricated from alendronate-conjugated carbon dots (Alen-CDs) and calcium-deficient hydroxyapatite (CDHA, the mineral component of bones) scaffolds is offered as a novel multifunctional BSM for in vivo osteoclasts deactivation and fluorescence imaging. The fluorescent Alen-CDs were hydrothermally prepared using phytic acid as carbon source, followed by conjugating alendronate, for controlled alendronate release and fluorescent imaging under acidic conditions. As-prepared fluorescent Alen-CDs were consecutively immobilized on surfaces of CDHA scaffolds, exhibiting high affinity by bisphosphonate group, easily fabricated from α-tricalcium phosphate (α-TCP) paste using three-dimensional (3D) printing system. The resultant Alen-CDs@CDHA caused a significant decrease (> 50%) in viability of osteoclasts at 7 days after in vitro treatment. Furthermore, when Alen-CDs@CDHA was implanted in balb/c nude mice for in vivo evaluation, we found Alen-CDs@CDHA to be suitable for bone imaging through fluorescence signals, without necrosis or inflammatory symptoms in the epidermal tissues. Thus, these observations offer new opportunities for a novel and revolutionary use of Alen-CDs@CDHA as highly specific multifunctional BSM for bone diagnosis and imaging, and as bone-specific drug delivery materials, eventually providing anti-osteoclastogenic treatments solution for degenerative bone disorders. STATEMENT OF SIGNIFICANCE: Alen-CDs@CDHA significantly reduced the viability of osteoclasts and fluorescently imaged in vivo after transplantation, releasing drug via pH modulation. The development of fluorescence materials for bone imaging remains still a major challenge in the biomedical field owing to the shortage of selectivity and specificity. The results could lead to improvements in bone treatment strategies, as it could reduce the invasiveness of procedures and the associated negative outcomes, and increase the precision of strategies. Further, we believe that this study will be of interest to the readership of your journal as clearly focuses on the advancement of a biomaterial, where we have engineered a substance to substitute bone and integrate with a living system.

Targeting Burkitt's Lymphoma Using Novel, Discrete Carbon Nanotubes

Burkitt's lymphoma is an incredibly aggressive B-cell derived non-Hodgkin lymphoma that is quickly fatal if left untreated. Doxorubicin (Doxo) provides one method of treatment for this cancer, but has numerous common side effects including significant weight loss, nausea, hair loss, bone marrow suppression, vomiting, rash, heart damage, allergic reaction, tissue damage at injection site, and treatment-related leukemia. Therefore, better therapies which deliver Doxo specifically to tumor cells hold great promise for a more effective therapy with reduced side-effect severity. Carbon nanotubes (CNTs) have long been of interest as drug delivery transporters because of their unique structure, high drug-binding surface area, and variety of customization options. Unfortunately, most CNTs have proven toxic during in vivo studies because of the presence of high residual contaminants and near-unavoidable aggregative properties. A novel form of clean, discrete CNT (dCNT) marketed as Medical Grade MOLECULAR REBAR® (MGMR®) lack the aggregative problems and high toxin levels of CNT before them and show promise as a drug delivery technology.We observed no toxicity of dCNTs in vitro across multiple cell types or in vivo in mice establishing a high MTD of at least 70 mg/kg by an i.v. route. Biodistribution studies of Lu-177 radioisotope labeled dCNTs (single tail vein injection into BALB/C female mice) demonstrated tolerability and clearance from various organs. Mice were sacrificed at 7 timepoints under 34d and many organs were analyzed for accumulation. The liver, spleen, and blood identified one route of clearance and distribution, but it was interesting to note that the bone retained ~10% of injected dCNTs at all time points, demonstrating potential for bone targeted therapies. Further targeting to the bone was possible by grafting commonly available bone chelators to the surface of the dCNT to create BT-dCNT as shown by significantly great adhesion (p<0.05) to ex vivo bone.Protocols were then developed to saturate the surface of the dCNT with therapeutic Active Pharmaceutical Ingredients (API) and then characterize their elution/release over time. We then optimized methods for drug-loading and observed a slow drug release profile for most drugs. Enhanced pharmacokinetic information was tested in cell culture and then confirmed by animal experimentation. In addition, cell penetration and delivery study of MGMR® in vitro showed that endocytosis is nontoxic and well tolerated and that small peptides can be delivered to cancer cells and retain their functionality. Zeta Potential measurements (-40 to -50 mV) and Dynamic Light Scattering validated the stability of different dCNTs over time.Lastly, in a Raji-Burkitt's lymphoma mouse model, we saw significant increases in survival, and decreases in tumor burden in mice treated with Doxo (free, in dCNTs, and in BT-dCNTs vs no drug control). SCID mice were inoculated via the tail vein with 2x105 Raji B. luc+ tumor cells and treated once a week with one of the following: Doxo (5 mg/kg), dCNTs (14.29 mg/kg), dCNTs (14.29 mg/kg) + Doxo (5 mg/kg), BT-dCNTs (14.29 mg/kg) + Doxo (5 mg/kg), or no treatment. Using whole body bioluminescence imaging (BLI), we saw that the groups receiving both the Doxo-loaded dCNTs and the Doxo-loaded BT-dCNTs had greatly reduced tumor burden over time compared to untreated groups, and a tumor burden similar to those that received free Doxo alone (Figure 1A). Both groups receiving Doxo-loaded dCNTs also showed increased survival compared to the Doxo alone group (Figure 1B). Importantly, the typical signs of Doxo toxicity in mice, characterized by rapid weight loss or drug-related mortality, were not seen in the groups that received Doxo-loaded CNTs (Figure 1C).In summary, dCNTs is a highly promising biomedical tool for drug delivery for clinical applications. Our dCNTs represent a promising platform for use in biomedical research as they have proven to be nontoxic, tunable, stable, and capable of drug delivery and organ-specific targeting. This novel technology gives great promise for the bone-targeting ability of novel carbon nanotubes in vivo as a bone-specific drug delivery mechanism for diseases that grow in bone or bone marrow specifically. [Display omitted] DisclosuresFalank:BioPact Ventures, LLC: Research Funding. Tassett:BioPact Ventures, LLC: Employment. Harris:BioPact Ventures, LLC: Research Funding. Farrell:BioPact Ventures, LLC: Research Funding. Everill:BioPact Ventures, LLC: Employment. Reagan:BioPact Ventures, LLC: Research Funding.

Biphasic composite of calcium phosphate-based mesoporous silica as a novel bone drug delivery system

We reported the new biphasic composites of calcium phosphate and mesoporous silica material (CaP@MSi) in the form of powders and pellets as a potential bone drug delivery system for doxycycline hydrochloride (DOX). The CaP@MSi powders were synthesized by cationic surfactant-templating method. The effects of 10, 20, and 30% CaP content in the CaP@MSi powders on the molecular surface structure, the cytotoxicity against osteoblast cells in vitro, and the mineralization potential in simulated body fluid were investigated. The CaP@MSi characterized by the highest mineralization potential (30% CaP content) were used for DOX adsorption and pelletization process. The CaP which precipitated in the CaP@MSi composites was characterized as calcium-deficient with the Ca:P molar ratio between 1.0 and 1.2. The cytotoxicity assays demonstrated that the CaP content in MSi increases osteoblasts viability indicating the CaP@MSi (30% CaP content) as the most biocompatible. The combination of CaP and MSi was an effective strategy to improve the mineralization potential of parent material. Upon immersion in simulated body fluid, the CaP of composite converted into the bone-like apatite. The obtained pellets preserved the mineralization potential of CaP@MSi and provided the prolonged 5-day DOX release. The obtained biphasic CaP@MSi composites seem to have an application potential as bone-specific drug delivery system.

Mesoporous silica-based hybrid materials for bone-specific drug delivery.

A mesoporous silica-based drug delivery device potentially useful for bone-specific drug delivery has been designed, developed and characterized starting from MSU-type mesoporous silica. The proposed system consists of a mesoporous silica nanoparticles (MSN) based vehicle, presenting alendronate as a targeting functionality for bone tissue while ibuprofen is used as a model molecule for the drugs to be delivered. The particles are functionalized on the external surface using a propionitrile derivative that is successively hydrolyzed to a carboxylic group. Alendronate, one of the most used member of the diphosphonate drug class, is electrostatically bonded to the external carboxyl functionalities of mesoporous silica. The obtained material has been characterized by powder X-ray diffraction, N2 adsorption–desorption porosimetry, UV-vis spectrophotometry, FT-IR spectrometry and MAS-NMR 13C and 29Si. Hydroxyapatite, which simulates the bone matrix, has been synthesized with the aim of testing the targeting activity of the obtained device. In a separate test, the MSNs have been loaded with ibuprofen, a non-steroidal anti-inflammatory drug (NSAID), and its release has been determined under neutral conditions by HPLC. Moreover, biological tests were carried out. The tested devices did not show any toxicity towards normal cells, confirming their high biocompatibility and the lack of off-target effects.

Multi-drug delivery system based on lipid membrane mimetic coated nano-hydroxyapatite formulations

Local delivery systems from an osteoconductive biomaterial are suggested as a promising strategy to avoid simultaneously peri-implant traumas and to induce tissue regeneration. In this work, it is detailed the design and construction of a multi-drug delivery formulation based on lipid membrane mimetic coated nano-hydroxyapatite, LMm/nano-HA, as a bone-specific drug delivery approach. The optimal LMm/nano-HA formulation was selected after analysing the lipid/nano-HA interaction by dynamic light scattering (DLS), ζ-potential, transmission electron microscopy (TEM), polarized optical microscopy (POM), differential scanning calorimetry (DSC) and UV–vis spectroscopy. After the initial screening, Ciprofloxacin and Ibuprofen simultaneous -load and -release efficiency from selected LMm/nano-HA was assessed. pH-responsive kinetic profiles of local drug distribution were characterized and compared with currently applied systemic doses. Finally, the systems’ biocompatibility and drug released activity were positively validated. The obtained results demonstrated that LMm/nano-HA formulations can represent a valuable multi-modal platform in bone tissue therapies.

Fracture-Targeted Delivery of β-Catenin Agonists via Peptide-Functionalized Nanoparticles Augments Fracture Healing.

Despite several decades of progress, bone-specific drug delivery is still a major challenge. Current bone-acting drugs require high-dose systemic administration which decreases therapeutic efficacy and increases off-target tissue effects. Here, a bone-targeted nanoparticle (NP) delivery system for a β-catenin agonist, 3-amino-6-(4-((4-methylpiperazin-1-yl)sulfonyl)phenyl)-N-(pyridin-3-yl)pyrazine-2-carboxamide, a glycogen synthase kinase 3 beta (GSK-3β) inhibitor, was developed to enhance fracture healing. The GSK-3β inhibitor loading capacity was found to be 15 wt % within highly stable poly(styrene-alt-maleic anhydride)-b-poly(styrene) NPs, resulting in ∼50 nm particles with ∼ -30 mV surface charge. A peptide with high affinity for tartrate-resistant acid phosphatase (TRAP), a protein deposited by osteoclasts on bone resorptive surfaces, was introduced to the NP corona to achieve preferential delivery to fractured bone. Targeted NPs showed improved pharmacokinetic profiles with greater accumulation at fractured bone, accompanied by significant uptake in regenerative cell types (mesenchymal stem cells (MSCs) and osteoblasts). MSCs treated with drug-loaded NPs in vitro exhibited 2-fold greater β-catenin signaling than free drug that was sustained for 5 days. To verify similar activity in vivo, TOPGAL reporter mice bearing fractures were treated with targeted GSK-3β inhibitor-loaded NPs. Robust β-galactosidase activity was observed in fracture callus and periosteum treated with targeted carriers versus controls, indicating potent β-catenin activation during the healing process. Enhanced bone formation and microarchitecture were observed in mice treated with GSK-3β inhibitor delivered via TRAP-binding peptide-targeted NPs. Specifically, increased bone bridging, ∼4-fold greater torsional rigidity, and greater volumes of newly deposited bone were observed 28 days after treatment, indicating expedited fracture healing.

ERRATUMRosuvastatin Implant for Local Bone Specific Drug Delivery in Osteoporotic Bone Fracture (Journal of Biomaterials and Tissue Engineering, Vol. 5(7), pp. 565–569 (2015))

ERRATUMRosuvastatin Implant for Local Bone Specific Drug Delivery in Osteoporotic Bone Fracture (Journal of Biomaterials and Tissue Engineering, Vol. 5(7), pp. 565–569 (2015))

Gelatin Nanoparticles with Enhanced Affinity for Calcium Phosphate.

Gelatin nanoparticles can be tuned with respect to their drug loading efficiency, degradation rate, and release kinetics, which renders these drug carriers highly suitable for a wide variety of biomedical applications. The ease of functionalization has rendered gelatin an interesting candidate material to introduce specific motifs for selective targeting to specific organs, but gelatin nanoparticles have not yet been modified to increase their affinity to mineralized tissue. By means of conjugating bone-targeting alendronate to biocompatible gelatin nanoparticles, a simple method is developed for the preparation of gelatin nanoparticles which exhibit strong affinity to mineralized surfaces. It has been shown that the degree of alendronate functionalization can be tuned by controlling the glutaraldehyde crosslinking density, the molar ratio between alendronate and glutaraldehyde, as well as the pH of the conjugation reaction. Moreover, it has been shown that the affinity of gelatin nanoparticles to calcium phosphate increases considerably upon functionalization with alendronate. In summary, gelatin nanoparticles have been developed, which exhibit great potential for use in bone-specific drug delivery and regenerative medicine.

Hydroxyapatite nanocrystals as a smart, pH sensitive, delivery system for kiteplatin.

Hydroxyapatite (HA) nanocrystals are important inorganic constituents of biological hard tissues in vertebrates and have been proposed as a bone substitute or a coating material for prostheses in biomedicine. Hydroxyapatite is also amenable for its capacity to bind to a great variety of biomolecules and therapeutic agents. As drug carriers, apatite nanoparticles also have the advantage of pH dependent solubility and low toxicity. Thus HA nanoparticles are negligibly soluble at physiological pH but their dissolution is accelerated at lower pH such as that typically found in the vicinity of tumors. In the present study we have investigated the adsorption on and the release from biomimetic HA nanoparticles of two platinum derivatives of cis-1,4-diaminocyclohexane ([PtX2(cis-1,4-DACH)], X2 = Cl2 (1) and 1,1-cyclobutanedicarboxylate (CBDCA, 2)). The first of the two compounds proved to be active against colon cancer cells also resistant to oxaliplatin. The release has been investigated as a function of pH to mimic the different physiological environments of healthy tissues and tumors, and the in vitro cytotoxicity of the releasates from the HA matrices has been assessed against various human cancer cell lines. The results fully confirmed the potential of 1-loaded HA nanoparticles as bone-specific drug delivery devices.

High mineral affinity of polyphosphoester ionomer–phospholipid vesicles

Bone-specific drug delivery is important for the treatment of osteoporosis and osseous metastases. However, there have been limitations in the design of drug carriers having bone affinity. We synthesized amphiphilic polyphosphoester ionomers (CH-PHE) and modified them to 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) vesicles. The ζ-potential of the vesicles was decreased by immobilization of CH-PHE; the amount was influenced by the structure and fraction of CH-PHE. The release rate of 5-carboxyfluorescein from the vesicles could be controlled by changing the fraction of DOPC and CH-PHE. In particular, the release of CF from DOPC vesicles containing 3% CH-PHE was most reduced. In addition, the enzymatic degradation of DOPC was reduced by immobilization with polyphosphoester ionomers; enzyme tolerance was increased with an increase in the molar fraction of polyphosphoester ionomers. Hemolytic activity of the phospholipid vesicles bearing CH-PHE was infrequently observed and was similar to that of the DOPC vesicles. Although a decrease in the viability of mouse osteoblastic cells (MC3T3-E1) in contact with the vesicles bearing CH-PHE was observed when the DOPC concentration of the vesicles bearing 20 mol % CH-PHE with highly ionized units was greater than 200 μM, the cytotoxicity was diminished by sodium salt formation of the CH-PHE. The affinity of the vesicles to calcium deposits generated by MC3T3-E1 cells was significantly improved by the immobilization polyphosphoesters.

Nanocrystalline carbonate-apatites: role of Ca/P ratio on the upload and release of anticancer platinum bisphosphonates

In the present study two nanocrystalline apatites have been investigated as bone-specific drug delivery devices to be used for treatment of bone tumors either by local implantation or by injection. In order to assess how the Ca/P ratio can influence the adsorption and release of anticancer platinum-bisphosphonate complexes, two kinds of apatite nanocrystals having different Ca/P ratios but similar morphologies, degree of crystallinity, and surface areas have been synthesized and characterized. The two platinum-bisphosphonate complexes considered were the bis-{ethylenediamineplatinum(ii)}-2-amino-1-hydroxyethane-1,1-diyl-bisphosphonate and the bis-{ethylenediamineplatinum(ii)}medronate. The Ca/P ratio plays an important role in the adsorption as well as in the release of the two drugs. In fact, the apatite with a higher Ca/P ratio showed greater affinity for both platinum complexes. Also the chemical structure of the two Pt complexes appreciably affects their affinity towards as well as their release from the two kinds of apatites. In particular, the platinum complex whose bisphosphonate contains a free aminic group showed greater upload and smaller release. The cytotoxicity of the Pt complexes released from the apatite was tested against human cervical, colon, and lung cancer cells as well as against osteosarcoma cells. In agreement with previous work, the Pt complexes released were found to be more cytotoxic than the unmodified complexes.

Smart delivery of antitumoral platinum complexes from biomimetic hydroxyapatite nanocrystals

This study widens the role of biomimetic hydroxyapatite (HA) nanocrystals as bone substitutes and describes how they can be used as bone-specific drug delivery devices for in situ treatment of bone tumors upon local implantation. The adsorption and release kinetics of bis-{ethylenediamineplatinum(II)}-2-amino-1-hydroxyethane-1,1-diyl-bisphosphonate and bis-{ethylenediamineplatinum(II)}medronate on two kinds of HA nanocrystals having different morphologies, crystallinity degrees and surface areas have been investigated. The different chemical structures of the two Pt complexes appreciably affect not only the affinity towards the two kinds of HA, but also their release. The Pt complex loading is slightly greater for the HA characterized by lower crystallinity and higher surface area, with respect to the more crystalline one. The cytotoxicity of Pt complexes released from the HA were tested against human cervix carcinoma cells and, interestingly, were found to be more cytotoxic than the unmodified complexes. The released Pt species are therefore the active dichloridoethylenediamineplatinum(II) or related solvato species formed by Pt-bisphosphonate bond breaking.

Adsorption and Conformational Change of Myoglobin on Biomimetic Hydroxyapatite Nanocrystals Functionalized with Alendronate

The chemical conjugation of bisphosphonates (BPs), specifically alendronate, to hydroxyapatite could be an effective means to impart to it fine-tuned bioactivity. Horse heart myoglobin (Mb), a well-characterized protein, has been adsorbed onto biomimetic hydroxyapatite nanocrystals (nHA) and onto the nHA/alendronate conjugate powdered samples. The obtained materials have potential use in bone implantation and as prospective drug-delivery devices. The kinetic absorption of Mb onto nHA is dramatically affected by its functionalization with alendronate. The covering of the nHA surface by alendronate inhibits the adsorption of myoglobin. The adsorption mechanisms of the protein were studied by spectroscopic techniques (UV-vis and surface-enhanced Raman spectroscopy). The results indicate that the protein changes conformation upon adsorption on the inorganic substrate. In particular, the interaction with nHA alters the coordination state of the iron in the heme through the formation of a hexacoordinated low-spin Mb heme, possibly involving the distal histidine. Instead, the covering of the nHA surface by alendronate does not adsorb the protein but preserves the coordination state of the heme moiety. This study could be of significance either in the field of biomaterials science, in particular, to fine tune a bone-specific drug delivery device and to test nHA as a new support for heterogeneous catalysis, improving the understating of enzyme immobilization.

Bone-specific drug delivery systems: approaches via chemical modification of bone-seeking agents.

Despite several decades of progress, bone-specific delivery is still limited by the unique anatomical features of bone, which mainly consists of inorganic hydroxyapatite. A practical approach to this problem is to produce targeted drugs that have a high affinity for hydroxyapatite. Bisphosphonates are a class of synthetic compounds structurally related to pyrophosphate. Bisphosphonates rapidly localise on the bone surface after being administered either intravenously or orally, since the P-C-P portion of the bisphosphonate structure has high affinity for hydroxyapatite. Therefore, bisphosphonate modification might be a promising method for targeting drugs selectively to the bone. Bisphosphonate-conjugated drugs are hydrophilic and highly water-soluble due to the acidic nature of the bisphosphonate moiety at physiological pH, and therefore they hardly permeate through the biological membrane of soft tissues. These physicochemical changes also reduce the intrinsic susceptibility of the drug to metabolism, promoting urinary or biliary excretion as unchanged drug. All these physicochemical and pharmacokinetic alterations contribute to the exceptional skeletal disposition of bisphosphonate-conjugated drugs. Bisphosphonate conjugation is based on chemical modification of the targeting molecule, and therapeutically optimised bisphosphonate derivatives have to be custom-developed on a case-by-case basis. The bisphosphonate moiety is usually coupled with the targeting drug through a specific linkage. The high affinity of bisphosphonate conjugates for the bone is not simply dependent on the bisphosphonate moiety but on the resultant molecule as a whole, including the linker and the linked drug. Lipophilicity (represented as log P) appears to be an appropriate index for predicting the osteotropic properties of bisphosphonate derivatives. Several strategies using bisphosphonate-conjugated drugs have been investigated at a laboratory level with the aim of obtaining therapeutically optimised treatments for conditions such as osteoporosis, osteoarthritis and bone cancer. In each case, the intention is to achieve prolonged local exposure to high concentrations of the targeting drug, thereby improving therapeutic index by enhancing pharmacological efficacy and minimising systemic adverse effects. Although most examples of bone-specific drug delivery via bone-seeking agents still remain in preclinical studies, several phosphonate-coupled radiopharmaceuticals, such as samarium-153 complexed to tetraphosphonate, are expected to be an effective pain palliation therapies for metastatic bone cancer and are currently being developed in clinical trials. Furthermore, recent reports on bisphosphonate-modified proteins have illustrated the feasibility of bone-specific delivery of biologically active protein drugs, such as cytokines and growth factors.