Concentrations Of Alendronate Research Articles

High throughput analysis of alendronate in human samples with derivatization-free hydrophilic-interactive chromatography mass spectrometry

A derivatization-free hydrophilic-interactive chromatography-mass spectrometry (HILIC-MS/MS) method was developed for quantifying low levels of alendronate in human plasma. Alendronate was separated and concentrated using calcium co-precipitation and analyzed by HILIC-MS/MS, requiring only a 300 μL plasma sample for each analysis. The method is simpler, safer, and more environmentally friendly than the conventional LC-MS/MS method that requires solid-phase extraction and derivatization steps during sample pretreatment. The method was validated for selectivity, linearity, precision, accuracy, extraction recovery, matrix effect and limit of quantification. The between-run precisions were no more than 7.1 % with accuracy ranging from − 1.7–6.3 %; extraction recovery was determined to be 85.3 %; while validation results indicated that the method was suitable for accurately quantifying alendronate concentrations in the range from 0.2 to 50 ng/mL. The approach was used successfully for high throughput analysis of alendronate in more than 3700 plasma samples from 120 subjects in a bioequivalence study.

Alendronate induces skeletal alterations in the chicken embryonic development model

Alendronate, a nitrogen-containing bisphosphonate, has reported long-term clinical success in the management of distinct bone-related conditions, particularly in the modulation of post-menopausal osteoporosis. Nonetheless, whether the inhibitory activity over osteoclastic cells' functionality is widely acknowledged, contradictory evidence arises from the assessment of alendronate activity over osteoblastic populations. This may be of particular relevance in situations in which bone formation exceeds bone resorption, with further emphasis on embryonic development, since alendronate can cross the placental barrier and alendronate-based therapies are being extended into women of reproductive age. Accordingly, the present study aims to assess the effects of alendronate, at distinct concentrations (1.5E−10M to 1.5E−7M) on bone tissue development, within a translational animal model – the embryonic chicken development model. Embryos, at the beginning of osteogenesis (day 7) were exposed to different alendronate concentrations for 4 days. Embryos were following characterized for skeletal development by histomorphometric analysis upon histochemical staining, microtomographic analysis, and gene expression assessment of genes related to osteoclastogenic/osteoclastic and osteoblastogenic/osteogenic differentiation, as well as to the immuno-inflammatory activation. The findings revealed that exposure to alendronate had a dose-dependent impact on skeletal growth and mineralization. This effect was evidenced by diminished bone volume and reduced bone surface parameters, with the 1.5E−7M concentration leading to a remarkable reduction of over 50%. Additionally, a decreased osteoclastogenic/osteoclastic gene expression was verified, associated with a diminished osteoblastogenic/osteogenic program – within the 30–50% range for 1.5E-7 M, supporting the diminished bone formation process. An increased inflammatory activation may contribute, at least in part, to the attained outcomes. Overall present findings suggest a negative influence of alendronate on the embryonic bone development process in a dose-dependent manner, highlighting the potential risk of alendronate use during embryonic development.

Novel insights into the pharmacological modulation of human periodontal ligament stem cells by the amino-bisphosphonate Alendronate

Alendronate (ALN) is a second-generation bisphosphonate widely used for osteoporosis and cancer-induced bone lesions. Many studies have confirmed a strong relationship between osteonecrosis of the jaws (ONJ) development and oral bisphosphonates, especially ALN, although the molecular mechanisms underlying this pathology have not yet been elucidated. The reduction in bone turnover and vascularization usually observed in ONJ are the result of ALN action on different cell types harboured in oral microenvironment, such as osteoclasts, endothelial cells, and periodontal ligament stem cells (PDLSCs). In this perspective, the present study aims to investigate the effects of different ALN concentrations (2 μM, 5 μM, 10 μM, 25 μM, 50 μM) on the phenotype and functional properties of human PDLSCs (hPDLSCs). hPDLSCs showed a decrease in cell viability (MTT assay) only when treated with ALN concentration of 10 μM or larger for 48 h and 72 h. Cell cycle analysis revealed a moderate increase in proportion of S-phase cells after exposure to low ALN concentration (2–5 μM), an effect that was reverted after exposure to 10–50 μM ALN. Conversely, cell death was evidenced via Annexin V/PI assay at very high concentration of ALN (50 μM) after 4 days of treatment. In addition, we explored whether the effects of ALN on hPDLSCs growth and survival can be mediated by its ability to modulate oxidative stress. To this, we quantified the intracellular ROS amount and lipid peroxidation by using DCF probe and Bodipy staining, respectively. Flow cytometry analysis showed that ALN induced a dose-dependent reduction of intracellular oxidative stress and lipid peroxidation upon treatment with low concentrations at both 48 h and 72 h. Increased levels of oxidative stress was reported at 50 μM ALN and was also confirmed via TEM analysis. Despite the stability of the cellular immunophenotype, hPDLSCs showed impaired mobility after ALN exposure. Chronic exposure (7–14 days) to ALN in the range of 2–10 μM significantly decreased the expression of the differentiation-related factors ALP, RUNX2, COLI, and OPN as well as the osteogenic ability of hPDLSCs compared with untreated cells. Conversely, higher doses were found to be neutral. Our findings indicated that the effects of ALN on hPDLSCs behavior are dose-dependent and suggest a role for oxidative stress in ALN-induced cell death that may lead to novel therapeutic approaches for ONJ.

Targeting a therapeutically relevant concentration of alendronate for in vitro studies on osteoblasts

Objective Bisphosphonates like alendronate mainly exert their effects on osteoclasts. However, osteoblasts are also affected, but exposed to a much lower concentration in vivo than the osteoclasts. Given that the effects are dose-dependent, the intention of the study was to identify a therapeutically relevant concentration of alendronate for in vitro studies on osteoblasts. Materials and methods Primary human osteoblasts were incubated with alendronate (5, 20 and 100 µM) for 1, 3, 7 and 14 days. Proliferation and viability were assessed, and the effects on cellular growth and function were evaluated by multianalyte profiling of selected proteins in cell culture media using the Luminex 200TM. Results The viability was not affected by any of the dosages. Exposure to 5 µM alendronate had a neutral effect on osteoblast proliferation, and on secretion of osteogenic and inflammatory markers, while enhancing synthesis of a marker of angiogenesis. 20 µM alendronate induced a decline in proliferation and affected angiogenic and osteogenic biomarkers adversely. 100 µM alendronate reduced proliferation dramatically, and this dosage was excluded from further experiments. Conclusion A concentration of 5 µM alendronate exerted effects on human osteoblasts that may translate to those observed in vivo and could therefore be relevant for in vitro studies.

Multiple Porous Synthetic Bone Graft Comprising EngineeredMicro-Channel for Drug Carrier and Bone Regeneration.

Due to high demand but limited supply, there has been an increase in the need to replace autologous bone grafts with alternatives that fulfill osteogenic requirements. In this study, two different types of bone grafts were tested for their drug carrying abilities along with their osteogenic properties. Two different types of alendronate-loaded bone grafts, Bio-Oss (bovine bone graft) and InRoad (biphasic synthetic bone graft) were observed to see how different concentrations of alendronate would affect the sustained release to enhance osteogenesis. In this study, defected ovariectomize-induced osteoporotic rat calvarias were observed for 28 days with three different concentrations of alendronate (0 mg, 1 mg, 5 mg) for both Bio-Oss and InRoad. A higher concentration (5 mg) allowed for a more controlled and sustained release throughout the 28-day comparison to those of lower concentrations (0 mg, 1 mg). When comparing Bio-Oss and InRoad through histology and Micro-CT, InRoad showed higher enhancement in osteogenesis. Through this study, it was observed that alendronate not only brings out robust osteogenesis with InRoad bone grafts, but also enhances bone regeneration in an alendronate-concentration-dependent manner. The combination of higher concentration of alendronate and multiple porous bone graft containing internal micro-channel structure of InRoad resulted in higher osteogenesis with a sustained release of alendronate.

The Effect of Alendronate on Osteoclastogenesis in Different Combinations of M-CSF and RANKL Growth Factors.

Bisphosphonates (BPs) are compounds resembling the pyrophosphate structure. BPs bind the mineral component of bones. During the bone resorption by osteoclasts, nitrogen-containing BPs are released and internalized, causing an inhibition of the mevalonate pathway. As a consequence, osteoclasts are unable to execute their function. Alendronate (ALN) is a bisphosphonate used to treat osteoporosis. Its administration could be associated with adverse effects. The purpose of this study is to evaluate four different ALN concentrations, ranging from 10−6 to 10−10 M, in the presence of different combinations of M-CSF and RANKL, to find out the effect of low ALN concentrations on osteoclastogenesis using rat and human peripheral blood mononuclear cells. The cytotoxic effect of ALN was evaluated based on metabolic activity and DNA concentration measurement. The alteration in osteoclastogenesis was assessed by the activity of carbonic anhydrase II (CA II), tartrate-resistant acid phosphatase staining, and actin ring formation. The ALN concentration of 10−6 M was cytotoxic. Low ALN concentrations of 10−8 and 10−10 M promoted proliferation, osteoclast-like cell formation, and CA II activity. The results indicated the induction of osteoclastogenesis with low ALN concentrations. However, when high doses of ALN were administered, their cytotoxic effect was demonstrated.

Sustained delivery of growth factors and alendronate using partially demineralized dentin matrix for endogenous periodontal regeneration

The decreasing number and reduced osteogenic differentiation capacity of mesenchymal stem cells (MSCs) and the excessive proliferation of osteoclasts in periodontal defects lead to difficulty in periodontal regeneration. Growth factors have powerful impacts on stem cell recruitment and differentiation, and alendronate (ALN) is a potent inhibitor of osteoclasts, which exert precise roles in periodontal regeneration. However, they are all hydrophilic molecules, typically delivered in a soluble format and rapidly released at supraphysiologic doses, which have side effects. In this study, we developed a simple and robust approach to the sustained release of bioactive molecules for endogenous periodontal regeneration. The release system was based on partially demineralized dentin matrix (PDD) in which growth factors were entrapped and hydroxyapatite was only distributed in the interior of the dentin tubules. ALN was then anchored inside PDD, after which PDD-ALN was obtained. PDD-ALN could stably release physiological concentrations of BMP-2, VEGFA, and ALN in a sustained manner. This delivery system exhibits three synergistic effects on bone microenvironment: i) PDD-ALN enhanced MSC migration and their osteogenic differentiation related to the BMP/Smad signaling pathway; ii) PDD-ALN inhibited the formation and function of osteoclasts related to the NF-κB, p38, and ERK1/2 signaling pathways; iii) PDD-ALN enhanced angiogenesis in umbilical vein endothelial cells related to the VEGFA/VEGFR2 signaling networks. PDD-ALN ultimately promoted periodontal regeneration in a rat model. This simple and low-cost technology provides a new idea for constructing an efficient delivery system and has promising prospects for the repair of defects in bone metabolic diseases.

Alendronate-functionalized poly(amido amine) cryogels of high-toughness for biomedical applications

Novel bisphosphonic acid-functionalized poly(amido amine) (PAA) macromers were synthesized through aza-Michael addition of sodium alendronate (ALE) and 5-amino-1-pentanol (AP) to N,N′-methylene bisacrylamide at two different molar ratios, with macromer only having AP as comparison control. The macromers were photopolymerized to cryogels whose swelling, biodegradation and mineralization properties were studied. Biodegradation was most strongly affected by the macromer molecular weight. In mineralization studies, the control cryogel nucleated HAP, the others another type of biological apatite; the extent of mineralization depending on ALE concentration. Cryogel-apatite composites were studied by SEM, FTIR, XRD and thermogravimetric analysis. Mechanical tests reveal compressions up to 97% for cryogels, showing their high toughness. Young's modulus and compressive fracture stress increase with ALE content to 10 kPa and 2.2 ± 0.4 MPa, respectively. The modulus increased significantly in mineralization due Ca2+-ALE bindings forming physical cross-links. Degradation products of ALE-containing cryogels showed dose, composition and cell type dependent cytotoxicity when incubated with osteosarcoma cells lines, Saos-2 and U-2 OS, and healthy C2C12 muscle cells.

In vitro effects of alendronate on fibroblasts of the human rotator cuff tendon

BackgroundBone mineral density of the humeral head is an independent determining factor for postoperative rotator cuff tendon healing. Bisphosphonates, which are commonly used to treat osteoporosis, have raised concerns regarding their relationships to osteonecrosis of the jaw and to atypical fracture of the femur. In view of the prevalence of rotator cuff tear in osteoporotic elderly people, it is important to determine whether bisphosphonates affect rotator cuff tendon healing. However, no studies have investigated bisphosphonates’ cytotoxicity to human rotator cuff tendon fibroblasts (HRFs) or bisphosphonates’ effects on rotator cuff tendon healing. The purpose of this study was to evaluate the cytotoxicity of alendronate (Ald), a bisphosphonate, and its effects on HRF wound healing.MethodsHRFs were obtained from human supraspinatus tendons, using primary cell cultures. The experimental groups were control, 0.1 μM Ald, 1 μM Ald, 10 μM Ald, and 100 μM Ald. Alendronate exposure was for 48 h, except during a cell viability analysis with durations from 1 day to 6 days. The experimental groups were evaluated for cell viability, cell cycle and cell proliferation, type of cell death, caspase activity, and wound-healing ability.ResultsThe following findings regarding the 100 μM Ald group contrasted with those for all the other experimental groups: a significantly lower rate of live cells (p < 0.01), a higher rate of subG1 population, a lower rate of Ki-67 positive cells, higher rates of apoptosis and necrosis, a higher number of cells with DNA fragmentation, higher caspase-3/7 activity (p < 0.001), and a higher number of caspase-3 positive staining cells. In scratch-wound healing analyses of all the experimental groups, all the wounds healed within 48 h, except in the 100 μM Ald group (p < 0.001).ConclusionsLow concentrations of alendronate appear to have little effect on HRF viability, proliferation, migration, and wound healing. However, high concentrations are significantly cytotoxic, impairing cellular proliferation, cellular migration, and wound healing in vitro.

Effect of photobiomodulation on the viability of osteoblasts and fibroblasts submitted to alendronate sodium or zoledronic acid

The goal of this study is to evaluate the effect of photobiomodulation therapy (PBMT) on the viability of osteoblasts and cultured fibroblasts in different concentrations of alendronate or zoledronic acid. Two cell lines: osteoblast-like mouse cells (OSTEO 1) and human buccal mucosa fibroblast (FMM1) were used. Cells were submitted to different concentrations of bisphosphonates (1 μM, 10 μM, and 100 μM sodium alendronate and 3 μM, 5 μM and 10 μM zoledronic acid) for 24 hours. Next, the cultures received PBMT. The irradiations were applied with a diode laser (InGaAIP, 660 nm, 30 mW, spot 0.028 cm2) in continuous, punctual and contact mode at two energy densities: 5 J/cm2 (4.5 s) or 10 Jcm2 (9s) with 6 hours-intervals. Cell viability was determined by mitochondrial activity assay (MTT) 24 h after the last irradiation. The data were compared by the one way- ANOVA, complemented by the Tukey’s test (p &lt; 0.05). Sodium alendronate at concentrations of 100 μM and 10 μM and zoledronic acid at 10 μM concentration showed higher long-term toxicity. The cellular viability of the PBMT treated group was significantly higher than that of the negative control group. The same occurred with the osteoblasts treated with the highest concentrations of the drug (5 and 10 μM), despite not reaching the cell viability of the positive control group, it presented greater viability than the negative control where the cells were not irradiated. In the groups submitted to zoledronic acid, positive controls presented greater cell viability. We concluded that under the parameters applied in this study, PBMT at an energy density of 5 J/cm2 was able to revert the toxicity of sodium alendronate applied at the higher concentrations in both cell types, whereas zoledronic acid toxicity, regardless of its concentrations, was not influenced by PBMT.

Effects of local application of alendronate on early healing of extraction socket in dogs.

The aim of the present study was to assess the effects of alendronate (ALN) on bone remodeling following tooth extraction in a dog model. For the study, fifteen male Beagles dogs of approximately 12months of age were used. Mesial roots of four mandibular premolars were endodontically treated, and the distal roots were removed. ALN concentrations of 0.5, 1, and 2mg/mL were topically applied for 15min, while a sterile saline was used as a negative control. After the healing period of 1, 2, and 8weeks, the samples were analyzed by micro-CT and histology. Treatment with ALN increased vertical distance between the lingual and the buccal crestal bones. While the ALN-treated sockets had preserved more lingual bone areas, control sockets showed better preservation of the buccal bone areas. ALN treatment resulted in more osteoid formation within the extraction sockets compared with the control. Higher bone volume was found in ALN groups than in the control at 2-week and 8-week healing periods, reaching the significant difference only for the extraction sockets pooled for the ALN treatment. Although ALN treatment could not prevent buccal bone resorption following tooth extraction in dogs, it proved beneficial for the preservation of the lingual bone and formation of new bone within the socket. There was no clear relation between the ALN dosages and the alterations within the extraction sockets. ALN affects bone remodeling of the extraction socket. The optimal concentration remains to be determined in future studies.

Elimination of intravenous alendronate by hemodialysis: A kinetic study.

The potential utility of intravenous alendronate for the treatment of osteoporosis in hemodialysis patients was recently reported. However, the pharmacokinetics of intravenous alendronate in patients on hemodialysis is not clear. Six hemodialysis patients (mean age, 80.5 years) with osteoporosis who had received intravenous alendronate prior to the study were enrolled. The participants received a 30-min infusion of 900-μg alendronate intravenously at the beginning of the dialysis session. The blood flow rate (Qb) and dialysate flow rate (Qd) were set at 200 mL/min and 500 mL/min, respectively. All patients used the same dialyzer (1.5-m2 polysulfone membrane). At the completion of administration, plasma and dialysate samples were collected, and alendronate concentrations were determined using metal-free high-performance liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS). The plasma arterial alendronate concentration was 150.9 ± 46.09 ng/mL. It decreased through the dialyzer to 76.1 ± 34.1 ng/mL (venous alendronate concentration). Mean alendronate clearance was 113.9 ± 25.6 mL/min. Mean alendronate removal by hemodialysis, measured by the difference in arterial-venous concentrations, was 51.8%. Fifty percent of intravenous alendronate was removed by hemodialysis, which is nearly equal to elimination of alendronate in patients with normal renal function. The elimination by hemodialysis would decrease the risk of excessive accumulation in bone. UMIN 000027182.

Alendronate Release from UHMWPE-Based Biomaterials in Relation to Particle Size of the GUR Powder for Manufacturing.

Ultra-high molecular weight polyethylene (UHMWPE) is widely used in endoprosthetics and has been the subject of countless studies. This project investigates the dependence of alendronate (AL) release on the molecular weight of the UHMWPE used (GUR1020 and GUR1050). A 0.5 wt% AL was added to the UHMWPE during the production of the moldings. In addition to the 14-day release tests, biocompatibility tests such as live dead assay, cell proliferation assay (WST) and Lactate dehydrogenase test (LDH) with MG-63 cells as well as a tensile test according to DIN EN ISO 527 were carried out. The released AL concentration was determined by HPLC. A continuous release of the AL was observed over the entire period of 2 weeks. In addition, a correlation between molar mass and AL release was demonstrated. The GUR1020 showed a release four times higher than the GUR1050. Both materials have no negative influence on the proliferation of MG-63 cells. This was also confirmed in the live/dead assay by the increase in cell count. No cytotoxicity was detected in the LDH test. The addition of 0.5 wt% AL increased the elongation at break for GUR1020 by 23% and for GUR1050 by 49%. It was demonstrated that the choice of UHMWPE has an influence on the release of AL. The particle size in particular has a strong influence on the release behavior.

Development of terbium-sensitized fluorescence method for the determination of alendronate in biological samples followed by magnetic solid-phase extraction

A new method based on combining terbium-sensitized fluorescence method and magnetic solid-phase extraction (MSPE) was developed for the quantification of alendronate (ALD) in biological samples. The fluorescence intensity of Tb3+-ALD complex was measured at 545 nm following elution from polymer coated magnetic nanoparticles surface. MNPs modified with PEG-b-P(IA-co-DMAEMA) block copolymer were synthesized and the effects of various parameters on MSPE efficiency and the fluorescence intensity were investigated. The dynamic assay range for ALD determination was obtained (0.04 to 0.8 mg/L) with a quantification limit of 0.033 mg/L. The accuracy of recovery, reproducibility, and sensitivity of the developed method suggested that it can be feasibly developed to a platform for monitoring the ALD concentration in human urine and plasma samples.

In vitro assessment of alendronate toxic and apoptotic effects on human dental pulp stem cells.

Objective(s):Osteonecrosis of the jaw, as an exposed necrotic bone in the oral cavity, is one of the adverse effects of bisphosphonates, which have an affinity for bone minerals. This study investigates the cytotoxic effects of alendronate (ALN) as a nitrogen-containing bisphosphonate, on human dental pulp stem cells (hDPSCs). Materials and Methods:The mesenchymal stem cells (MSCs), obtained from third molar tooth pulps were characterized by immunophenotyping assay in order to identify surface markers to evaluate their expression. To detect multipotency hDPSCs, they were differentiate into osteocytes and adipocytes. Cell proliferation was measured by MTT assay. PI staining of DNA fragmentation by flowcytometry (sub-G1 peak) was performed for determination of apoptotic cells and Bax, Bcl-2, and cleaved caspase 3 expressions. Protein expression was detected by Western blotting. Results:As the results revealed, ALN decreased viable cells (in 0.8–100 µM) after 72 hr and 168 hr (P<0.001), significantly. ALN could lower cell proliferation in hDPSCs in a concentration and time-dependent manner. Sub-G1 peak as an indicator of flowcytometry histogram of treated cells by ALN, showed apoptosis was involved in ALN-induced cytotoxicity. Expressions of cleaved caspase 3 and Bax protein, as pro-apoptotic proteins, were increased and Bcl-2 protein as anti-apoptotic protein was decreased in response to increases in the concentration of ALN (0.8–25 µM).Conclusion:Long-term effects of ALN on cell proliferation and apoptosis in hDPSCs can result in either initiation or potentiation of ALN-induced osteonecrosis.

Alendronate delivery on amino modified mesoporous bioactive glass scaffolds to enhance bone regeneration in osteoporosis rats

The regeneration capacity of osteoporotic bones is generally lower than that of normal bones. Nowadays, alendronate (AL) are orally administrated for osteoporosis due to the inhibition of bone resorption. However, systemic administration of AL is characterized by extremely low bioavailability and high toxicity. In this study, the amino-modified mesoporous bioactive glass scaffolds (N-MBGS) were fabricated by a simple powder processing technique as a novel drug-delivery system for AL. The effects of AL on the osteogenic differentiation of bone mesenchymal stem cells derived from ovariectomized rats (rBMSCs-OVX) were first estimated. The loading efficiency and release kinetics of AL on N-MBGS were investigated in vitro and the osteogenesis of AL-loaded N-MBGS in rat calvarial defect model was detected by micro-CT measurements and the histological assay. Our results revealed that proper concentration of AL significantly promoted osteogenic differentiation of rBMSCs-OVX. The amount and delivery rate of AL were greatly improved through amino modification. Additionally, scaffolds with AL showed better bone formation in vivo, especially for the N-MBGS group. Our results suggest that the novel amino-modified MBGS are promising drug-delivery system for osteoporotic bone defect repairing or regeneration.The experimental schematic of the novel amino-modified MBGS as a promising drug-delivery system for osteoporotic bone regeneration.

High payload nanostructured lipid carriers fabricated with alendronate/polyethyleneimine ion complexes

Oral bioavailability of the anti-osteoporotic drug alendronate (AL) is limited to ≤ 1% due to unfavorable physicochemical properties. To augment absorption across the gastrointestinal mucosa, an ion pair complex between AL and polyethyleneimine (PEI) was formed and incorporated into nanostructured lipid carriers (NLCs) using a modified solvent injection method. When compared to free AL, ion pairing with PEI increased drug encapsulation efficiency in NLCs from 10% to 87%. Drug release from NLCs measured in vitro using fasted state simulated intestinal fluid, pH 6.5 (FaSSIF-V2) was significantly delayed after PEI complexation. Stability of AL/PEI was pH-dependent resulting in 10-fold faster dissociation of AL in FaSSIF-V2 than measured at pH 7.4. Intestinal permeation properties estimated in vitro across Caco-2 cell monolayers revealed a 3-fold greater flux of AL encapsulated as hydrophobic ion complex in NLCs when compared to AL solution (Papp = 8.43 ± 0.14 × 10−6 cm/s and vs. 2.76 ± 0.42 × 10−6 cm/s). Cellular safety of AL/PEI-containing NLCs was demonstrated up to an equivalent AL concentration of 2.5 mM. These results suggest that encapsulation of AL/PEI in NLCs appears a viable drug delivery strategy for augmenting oral bioavailability of this clinically relevant bisphosphonate drug and, simultaneously, increase gastrointestinal safety.

Alendronate-loaded, biodegradable smart hydrogel: a promising injectable depot formulation for osteoporosis

Alendronate (ALN) is a BCS III bone resorption inhibitor, with very poor oral bioavailability. Our approach is to develop a minimally invasive thermogelling system for prolonged local delivery of ALN. For this, different chitosan-based thermogels were developed and characterised in terms of gelation time, injectability, pH, viscosity and thermoreversibility. Chitosan/β-glycerophosphate (CS/βGP) hydrogel pursued temperature-dependent, thermoreversible gelation behaviour and was thus selected for drug loading. Increasing ALN concentration resulted in hydrogels with lower porosity and higher density. FTIR and DSC proved interaction between ALN, CS with βGP. CS/βGP hydrogel ensured controlled ALN release over 45–65 days depending on initial ALN loading. Freeze drying improved the shelf-life stability with minor impact on thermogelling character. In vivo injection of plain and ALN-loaded hydrogel in rats rapidly gelled 15 min post-injection. Based on histological examination, ALN-loaded thermogel showed less inflammatory response, faster proliferation and maturation of granulation tissue relative to plain thermogel. Hydrogels excised 21-days post-injection proved the biocompatibility and biodegradability of the system. The presented chitosan-based thermogel has significant positive attributes for site-specific, time-controlled, intra-articular delivery of ALN.

Would Be Prophylactic Administrations of Low Concentration of Alendronate an Alternative for Improving the Craniofacial Bone Repair? A Preliminary Study Focused in the Period of Cellular Differentiation and Tissue Organization.

Alendronate (ALN) is a nitrogen-bisphosphonate that may induce an anabolic effect on craniofacial bone repair when administrated in low doses. Based on this premise, this study analyzed the influence of prophylactic low doses of ALN on bone healing in defects created in rabbit mandible. A 5 × 2-mm diameter deep defect was created in the calvaria of 28 rabbits. Fourteen of these rabbits received previously 50 μg/kg of 1% sodium ALN for 4 weeks, while the other rabbits received only 0.9% physiological saline solution (control). Animals were euthanized at 15 and 60 days postsurgery (n = 7), and the data were analyzed using histomorphometry and immunohistochemistry using the anti-CD34, bone morphogenetic protein -2 (BMP-2), and transforming growth factor (TGF)-β1 antibodies. On the 15th day postsurgery, the specimens that received previous treatment with ALN demonstrated large vascular lumen and intense positivity to CD34 either concentrated in endothelium or cells spread among the reparative tissue. These results coincided with intense positivity for BMP-2+ cells and TGF-β1 that was concentrated in both cells and perivascular area. In contrast, the control group revealed scarce cells that exhibited CD34, BMP-2+, and the TGF-β1 was restricted for perivascular area on well-formed granulation tissue. These patterns of immunohistochemical result, especially found on the 15th day of analysis, seem to be responsible for the development of larger quantities of bone matrix in the specimens that receive ALN on the 60th day postsurgery. These preliminary results showed that the prophylactic administration of low doses of ALN might be an alternative to craniofacial bone craniofacial bone repair because it increases the immunopositivity for TGF-β1 and consequently improves the CD34+ and BMP-2+ cells on reparative sites.

Efficient Transdermal Delivery of Alendronate, a Nitrogen-Containing Bisphosphonate, Using Tip-Loaded Self-Dissolving Microneedle Arrays for the Treatment of Osteoporosis.

To improve the transdermal bioavailability and safety of alendronate (ALN), a nitrogen-containing bisphosphonate, we developed self-dissolving microneedle arrays (MNs), in which ALN is loaded only at the tip portion of micron-scale needles by a dip-coating method (ALN(TIP)–MN). We observed micron-scale pores in rat skin just after application of ALN(TIP)–MN, indicating that transdermal pathways for ALN were created by MN. ALN was rapidly released from the tip of MNs as observed in an in vitro release study. The tip portions of MNs completely dissolved in the rat skin within 5 min after application in vivo. After application of ALN(TIP)–MN in mice, the plasma concentration of ALN rapidly increased, and the bioavailability of ALN was approximately 96%. In addition, the decrease in growth plate was effectively suppressed by this efficient delivery of ALN in a rat model of osteoporosis. Furthermore, no skin irritation was observed after application of ALN(TIP)–MN and subcutaneous injection of ALN, while mild skin irritation was induced by whole-ALN-loaded MN (ALN–MN)—in which ALN is contained in the whole of the micron-scale needles fabricated from hyaluronic acid—and intradermal injection of ALN. These findings indicate that ALN(TIP)–MN is a promising transdermal formulation for the treatment of osteoporosis without skin irritation.