Epigenetic regulation of the miR142-3p/interleukin-6 circuit in glioblastoma.

Epigenetic regulation plays a critical role in glioblastoma (GBM) tumorigenesis. However, how microRNAs (miRNAs) and cytokines cooperate to regulate GBM tumor progression is still unclear. Here, we show that interleukin-6 (IL-6) inhibits miR142-3p expression and promotes GBM propagation by inducing DNA methyltransferase 1-mediated hypermethylation of the miR142-3p promoter. Interestingly, miR142-3p also suppresses IL-6 secretion by targeting the 3' UTR of IL-6. In addition, miR142-3p also targets the 3' UTR and suppresses the expression of high-mobility group AT-hook 2 (HMGA2), leading to inhibition of Sox2-related stemness. We further show that HMGA2 enhances Sox2 expression by directly binding to the Sox2 promoter. Clinically, GBM patients whose tumors present upregulated IL-6, HMGA2, and Sox2 protein expressions and hypermethylated miR142-3p promoter also demonstrate poor survival outcome. Orthotopic delivery of miR142-3p blocks IL-6/HMGA2/Sox2 expression and suppresses stem-like properties in GBM-xenotransplanted mice. Collectively, we discovered an IL-6/miR142-3p feedback-loop-dependent regulation of GBM malignancy that could be a potential therapeutic target.

Demethoxycurcumin-Loaded Chitosan Nanoparticle Downregulates DNA Repair Pathway to Improve Cisplatin-Induced Apoptosis in Non-Small Cell Lung Cancer.

Demethoxycurcumin (DMC), through a self-assembled amphiphilic carbomethyl-hexanoyl chitosan (CHC) nanomatrix has been successfully developed and used as a therapeutic approach to inhibit cisplatin-induced drug resistance by suppressing excision repair cross-complementary 1 (ERCC1) in non-small cell lung carcinoma cells (NSCLC). Previously, DMC significantly inhibited on-target cisplatin resistance protein, ERCC1, via PI3K-Akt-snail pathways in NSCLC. However, low water solubility and bioavailability of DMC causes systemic elimination and prevents its clinical application. To increase its bioavailability and targeting capacity toward cancer cells, a DMC-polyvinylpyrrolidone core phase was prepared, followed by encapsulating in a CHC shell to form a DMC-loaded core-shell hydrogel nanoparticles (DMC-CHC NPs). We aimed to understand whether DMC-CHC NPs efficiently potentiate cisplatin-induced apoptosis through downregulation of ERCC1 in NSCLC. DMC-CHC NPs displayed good cellular uptake efficiency. Dissolved in water, DMC-CHC NPs showed comparable cytotoxic potency with free DMC (dissolved in DMSO). A sulforhodamine B (SRB) assay indicated that DMC-CHC NPs significantly increased cisplatin-induced cytotoxicity by highly efficient intracellular delivery of the encapsulated DMC. A combination of DMC-CHC NPs and cisplatin significantly inhibited on-target cisplatin resistance protein, ERCC1, via the PI3K-Akt pathway. Also, this combination treatment markedly increased the post-target cisplatin resistance pathway including bax, and cytochrome c expressions. Thymidine phosphorylase (TP), a main role of the pyrimidine salvage pathway, was also highly inhibited by the combination treatment. The results suggested that enhancement of the cytotoxicity to cisplatin via administration of DMC-CHC NPs was mediated by down-regulation of the expression of TP, and ERCC1, regulated via the PI3K-Akt pathway.

Demethoxycurcumin-carrying chitosan-antibody core-shell nanoparticles with multitherapeutic efficacy toward malignant A549 lung tumor: from in vitro characterization to in vivo evaluation.

Targeting controlled release core-shell nanocarriers with the potential to overcome multidrug resistant (MDR) lung cancer were prepared based on demethoxycurcumin (DMC) loaded amphiphilic chitosan nanoparticles coated with an anti-EGFR antibody layer. The nanocarriers were characterized with regard to size with dynamic light scattering, SEM, and TEM. The characterization confirmed the nanocarriers to have a surface coating of the anti-EGFR antibody and a final size excellently suited for circulating targeting nanocarriers, i.e., <200 nm in diameter. In vitro drug release revealed extended quasi-Fickian release from the nanocarriers, with the anti-EGFR layer further reducing the release rate. Cell culture experiments using normoxic and MDR hypoxic cells overexpressing EGFR confirmed improved DMC delivery for anti-EGFR coated particles and revealed that the DMC was delivered to the cytoplasmic region of the cells, forming nanoprecipitates in lysosomes and endosomes. The effective endocytosis and targeting of the core-shell nanoparticles resulted in the nanocarriers achieving high cytotoxicity also against MDR cells. The therapeutic potential was further confirmed in an A549 xenograft lung tumor mouse model, where DMC loaded core-shell nanocarriers achieved about 8-fold reduction in tumor volume compared with control group over the 8 weeks of the investigation. Both in vitro and in vivo data suggest the anti-EGFR coated core-shell nanocarriers as highly promising for treatment of hypoxic MDR cancers, especially for non-small cell lung cancer.

Forming of demethoxycurcumin nanocrystallite-chitosan nanocarrier for controlled low dose cellular release for inhibition of the migration of vascular smooth muscle cells.

We report an efficient therapeutic approach to inhibit the migration and growth of vascular smooth muscle cells (VSMCs) via a low-dose sustained elution of a water-insoluble drug, demethoxycurcumin (DMC), through a self-assembled amphiphilic carbomethyl-hexanol chitosan (CHC) nanomatrix. Manipulating the cellular internalization and controlled cytotoxic effect of DMC-CHC nanoparticles over the VSMCs was elucidated. The DMC-CHC nanoparticles, which were systematically characterized in terms of structural morphology, surface potential, encapsulation efficiency, and DMC nanocrystallite distribution, exhibited rapid cellular uptake efficiency and considerably improved cytotoxic potency by 2.8 times compared to the free DMC. Under a cytotoxic evaluation, an improved antiproliferative effect and effective inhibition of VSMC migration as a result of highly efficient intracellular delivery of the encapsulated DMC in comparison to free DMC was achieved, which also was confirmed with a subsequent protein analysis. Cellular drug release and distribution of DMC after internalization into VSMCs was experimentally determined. This work may open a potential intracellular medicinal strategy with improved biological and therapeutic efficacy using the DMC-CHC nanoparticles illustrated in this work.

Use of irinotecan for treatment of small cell carcinoma of the prostate.

BACKGROUND: Prostatic small cell carcinoma (SCC) is a rare variant of prostate cancer. It is extremely aggressive and resistant to available therapies with a median survival range of 5-17 months. No standard chemotherapeutic regimen has been established for its treatment. In search of a new therapeutic approach, we examined the response of patient-derived prostatic SCC tissue xenografts to irinotecan, a topoisomerase I inhibitor. METHODS: A tumor tissue line was established from a patient's prostatic SCC by subrenal capsule grafting using NOD-SCID mice. Mice carrying subcutaneous transplants of the tumor line were then treated for 2 weeks with irinotecan alone and in combination with cisplatin. The effect on tumor volume, histopathology, and apoptosis were determined. RESULTS: The prostatic SCC tissue line resembled the donor tissue in morphologic and immunohistochemical features. Irinotecan (20 mg/kg/day; days 1-3, 8-10) completely arrested xenograft growth with a small reduction in tumor volume and only minor weight loss of the hosts (7%); irinotecan (12 mg/kg; same schedule) + cisplatin (2.5 mg/kg/day; days 1 and 8) had a similar effect, but with lower weight loss. While the growth inhibition involved apoptosis, it was also associated with a marked increase in autophagy. CONCLUSIONS: Tumor tissue lines established via subrenal capsule xenografting provide models with clinical relevance and the present study suggests that irinotecan could be useful for therapy of refractory prostatic SCC, in particular in combination with cisplatin.

Magnolol-loaded core-shell hydrogel nanoparticles: drug release, intracellular uptake, and controlled cytotoxicity for the inhibition of migration of vascular smooth muscle cells.

Encapsulation and release behavior of a water-insoluble drug, magnolol, using a core-shell polysaccharide-based nanoparticle, manipulating the cellular internalization and controlled cytotoxic effect of magnolol-loaded nanoparticles over the A10 vascular smooth muscle cells (VSMCs) was reported. A magnolol-polyvinylpyrrolidone (PVP) core phase was prepared, followed encapsulating by an amphiphilic carboxymethyl-hexanoyl chitosan (CHC) shell to form a magnolol-loaded core-shell hydrogel nanoparticles (termed magnolol-CHC nanoparticles). The resulting magnolol-CHC nanoparticles were employed for evaluation of drug release and controlled cytotoxic inhibition of VSMCs migration in vitro. A sustained release of the magnolol from the nanoparticles was determined. The magnolol-CHC nanoparticles exhibited outstanding cellular uptake efficiency, and under a cytotoxic evaluation, an increased antiproliferative effect and effective inhibition of VSMC migration as a result of efficient intracellular delivery of the encapsulated magnolol in comparison to free magnolol was achieved. We then envision a potential intracellular medication strategy with improved biological and therapeutic efficacy using the magnolol-CHC nanoparticles illustrated in this work.

Characterization and structure evolution of Ca-Al-CO3 hydrotalcite film for high temperature CO2 adsorption.

In this work, monodispersed layered double hydroxide (Ca-Al LDHs) nanoparticles were synthesized by hydrothermal coprecipitation. Uniform thin films of layered double hydroxide on porous anodic aluminum oxide (AAO) substrates were formed by a direct precipitation process in a homogeneous suspension containing monodispersed Ca-Al layered double hydroxide nanoparticles. It was found that the formation of a designed hydrotalcite-like phase is strongly dependent on the [Ca(2+)]/[Al(3+)] ratios, and that a minor CaCO3 phase could possibly form simultaneously, which is attributed to the greater insolubility of CaCO3 and the incompatibility of the ionic size of Al and Ca. The optimal CO2 adsorption capacity appears in the layered Ca-OH-Al structure with the composition ratio of 3:1. Furthermore, the CO2 adsorption mechanism varies with treatment temperature. Below 400 degrees C, the CO2 adsorption is attributed to the LDH structure with a large surface area and pore volume, but above that the adsorption is due to the formation of CaCO3 and CaO. The permeation behavior and CO2 absorption can be explained by a preferable chemical and physical absorption of CO2 on the layered double hydroxide and porous structure of the membrane.

Self-assembly behavior and doxorubicin-loading capacity of acylated carboxymethyl chitosans.

A new type of acylated carboxymethyl amphiphilic chitosan (ACC) with the use of acyl chain of varying lengths, from C(2) to C(12), and various degrees of acyl substitution was successfully synthesized and has been characterized in terms of its self-assembly behavior, structural stability, and drug encapsulation. The resulting nanostructure of the ACC nanoaggregates can be well manipulated through a control of hydrophobicity. Structural evolution of the self-assembled nanoaggregates is extensively characterized via (1)H NMR, FTIR, DSC, and TEM. A critical value of the hydrophobic effect, (X(DH) x X(Cn)), i.e., a product of "degree of acyl substitution" and "carbon number of acyl chain", can be employed as an indicator for structural variation of the nanoaggregates: when (X(DH) x X(Cn)) exceeded 1.5, the architecture of the nanoaggregates underwent a structural transformation from solid nanoparticle to hollow nanocapsules. The nanoaggregates exhibited an excellent colloidal and structural stability in aqueous medium. An improved affinity toward drug encapsulation, i.e., doxorubicin, can be technically designed according to the amphiphilic nature of the resulting nanoaggregates for drug delivery.

A nanoporous optically transparent CdS-pHEMA hybrid with high optical sensing capability to dielectric liquids.

A hydrogel-based functional hybrid with highly uniformly dispersed nanoparticulate CdS semiconductors is proposed. The hybrid is synthesized using an in situ polymerization following an in situ chemical reduction, where the resulting particle size and the distribution of CdS nanocrystals (NCs) can be narrowly manipulated. The hybrids, containing a relatively small amount of the CdS NCs, exhibit a pronounced photoluminescence spectrum shift when in contact with a number of dielectric liquids and such a pronounced dielectric-confinement effect has been experimentally verified and modeled in this study. The sensing capability of the hybrids with respect to dielectric liquids or molecules can be optically characterized and varied depending upon the intensity of the dielectric environment surrounding the hybrids. This work suggests that the transparent, nanoporous CdS-pHEMA hybrids can be used as highly efficient optical sensing materials.

Electrochemical behavior of nano Cu/poly(2-hydroxyethyl methacrylate) composite.

A novel electro-activated Cu/pHEMA nanocomposite was fabricated by coupling nano copper particles with poly(2-hydroxyethyl methacrylate) (pHEMA) matrix. The nano-sized copper particles ranging from 5 nm to 10 nm were in-situ synthesized in the presence of three-dimensional polymer matrixes by photo-polymerization of the copper ions with the HEMA monomer and followed by a strong reducing reaction. The electrochemical behaviors of the Cu/pHEMA nanocomposite were charcterized by cyclic voltammetry and alternating current impedance method. The results showed that the redox behavior of Cu/pHEMA nanocomposite was controlled by the crystallinity of copper particles and the interaction between copper and pHEMA.

Electrical-sensitive nanoparticle composed of chitosan and tetraethyl orthosilicate for controlled drug release.

In the present study, the nanoparticles with size of 50-130 nm composed of chitosan (CS) and tetraethyl orthosilicate (TEOS), which showed electrically-controlled release profile, were prepared. Myoglobin was encapsulated into the nanoparticles with an encapsulation efficiency as high as 91.5% and a maximum loading capacity of 70.2% that can be achieved. Here, TEOS was hydrolyzed and condensed to form interpenetration network with CS. Increase of TEOS content results in a shift of the release profile from swelling-controlled towards diffusion-controlled mechanism. The "slow-to-burst" and "near-zero" release of the myoglobin can be effectively controlled by applying DC electric field between "on" and "off".

Stimuli-responsive controlled drug release from magnetic-sensitive silica nanospheres.

A novel method for control burst releasing of drug via a high frequency magnetic field (HFMF) from magnetic-sensitive silica nanospheres was developed. The nanospheres were synthesized by a combination of emulsion and sol-gel process with the particles controlled at about 80 nm in diameter. Under repeated exposures to the high frequency magnetic stimulus, the drug release behaviors showed reproducible slow-to-burst profiles while consecutively applying the magnetic stimulus at 10-min switching time and the release profile restored immediately when the stimulus was removed. By taking this non-contact control-burst method, the magnetic silica nanospheres can be designed to treat the cancer therapy and urgent physiological needs.

A new type of gadodiamide-conjugated amphiphilic chitosan nanoparticle and its use for MR imaging with significantly enhanced contrastability.

Magnetic resonance imaging (MRI) has been one of the most frequently-used diagnostic tools with high dimensional precision and positioning accuracy in clinical practices. To achieve contrast enhancement, utilization of high-efficient MR imaging contrast agents becomes a prime consideration and is indispensably reinforced the diagnosis precision, especially for the emerging precision medicine. Gadolinium (Gd)-based complexes has been widely used in current clinical MRI operations, however, numerous side effects were reported and highlighted in clinic. Those drawbacks render specific unmet needs to be clinically and technically improved with a new version of Gd-based compound. Here we report a newly-synthesized amphiphilic Gadodiamide-conjugated carboxymethyl-hexanoyl chitosan (termed as CHC-Gd) hybrid. The gadodiamide was selected is due to its smallest molecular size among other Gd-based complexes reported in literature, which assumed to give least influence on the resulting physicochemical properties such as colloidal stability, nanostructural evolution, and cytocompability, particularly self-assembly capability, of the resulting hybrid upon practical uses. Experimental outcomes showed a successful synthesis of the CHC-Gd hybrid using a one-pot synthesis protocol, where the gadodiamide complexes were covalently attached to the carboxyl groups along the CHC backbone. Self-assembly behavior can be observed to form a sphere-like nanoparticle of 100-200 nm in size as of amphiphilic native CHC macromolecule. Experimental outcomes indicated a largely improved cytocompatibility of the hybrid, compared with free Gd, suggesting the Gd+3 ions were well stabilized in the CHC nanostructure. Excellent contrastability in-vitro and in particular in vivo were measured, where for in-vivo test, a 10-40-folded reduction in dosage, compared with clinical Gd dose, was used and demonstrated a comparative-to-better imaging resolution and brightness. Therefore, from this preliminary investigation, a potential translation to clinical practice through the use of newly-synthesized amphiphilic CHC-Gd hybrid appears to be relatively promising.

Self-assembled amphiphilic chitosan: A time-dependent nanostructural evolution and associated drug encapsulation/elution mechanism.

This investigation reports the nanostructural evolution and associated encapsulation and elution of a hydrophobic drug, demethoxycurcumin (DMC), as a molecular probe, with the carboxymethyl-hexanoyl chitosan (CHC), which has been a technically interesting amphiphilic chitosan-based polymer successfully developed in this lab for years. The self-assembly nature of the CHC in neutral aqueous solutions allowed efficient encapsulation of various drugs without deteriorating or changing drugs' activity. However, its self-assembly behavior associated with nanostructural stability or variation, in terms of residence time in aqueous solution has not been well characterized and how the CHC nanostructure may be altered upon entrapping a drug, followed releasing out of the nanostructure. In this study, the CHC/DMC assembled model was used to evaluate entrapping efficiency, CHC-DMC interaction, and nanostructural variation while the drug being encapsulated and released from the CHC nanoparticles. Experimental outcomes showed a fractal transition between nanoparticulate and short fiber-like network evolution of the CHC as time elapsed, with the presence or absence of the DMC probe. This entrapment of DMC is relatively efficient upon CHC assembly and the associated DMC arrangement inside the helical CHC macromolecule gave largely increasing space over the resulting CHC/DMC assembly. Its excellent colloidal and nanostructural stability over a reasonably long period of time in testing environment suggests that this CHC/DMC assembly not only provides a crucial advantage for drug delivery application but also considers as a nanostructural model for better understanding of the mechanism upon drug encapsulation and elution which may be applicable to alternative amphiphilic polysaccharide-based macromolecules.

Core/Single-Crystal-Shell Nanospheres for Controlled Drug Release via a Magnetically Triggered Rupturing Mechanism.

Core/single-crystal-shell nanospheres are constructed from a poly-(N-vinyl- 2-pyrrolidone) (PVP)-modified silica core with an outer layer of single-crystal iron oxide shell. The nanospheres show outstanding release-and-zero-release characteristics via the addition and removal, respectively, of an external high-frequency magnetic field.

Drug release behavior of chitosan-montmorillonite nanocomposite hydrogels following electrostimulation.

Nanocomposites hydrogel (nanohydrogel) composed of chitosan (CS) and montmorillonite (MMT) were prepared and systematically studied for drug release behavior following electrostimulation. The deterioration of the responsiveness and reversibility of CS upon repeated on-off electrostimulation switching operations are major limitations for clinical applications, as it suffers from too much structural instability for the precise control of the release of drug upon cyclic electrostimulation. To overcome these limitations, an inorganic phase, MMT, was incorporated in the CS matrix to enhance the anti-fatigue property and corresponding long-term stable release kinetics. X-ray diffraction analysis and time-dependent optical absorbance showed that the MMT incorporated into the nanohydrogel exhibited an exfoliated nanostructure. The exfoliated silica nanosheets are able to act as cross-linkers to form a network structure between the CS and MMT, and this difference in the cross-linking density strongly affects the release of vitamin B(12) under electrostimulation. With a lower MMT concentration (1 wt.%), the release kinetics of vitamin B(12) from the nanohydrogel shows a pseudo-zero-order release, and the release mechanism was changed from a diffusion-controlled mode to a swelling-controlled mode under electrostimulation. Further increasing the MMT content reduced both the diffusion exponent n and the responsiveness of the nanohydrogel to electrostimulation. In addition, a consecutively repeated "on" and "off" operation shows that the electroresponsiveness of the nanohydrogel with higher MMT concentrations was reduced, but its anti-fatigue behavior was considerably improved. In this work, the nanohydrogel with 2 wt.% MMT achieved a mechanically reliable and practically desirable pulsatile release profile and excellent anti-fatigue behavior, compared with that of the pure CS.

Effect of clay content on electrostimulus deformation and volume recovery behavior of a clay-chitosan hybrid composite.

Electrostimulus-responsive hybrid composites composed of chitosan (CS) and clay were successfully developed and systematically characterized. The addition of negatively charged clay as an ionic cross-linker strongly affect the cross-linking density as well as the mechanical property, swelling-deswelling behavior and fatigue property of the hybrids. With lower clay content, the crystallinity of the CS was slightly reduced, resulting in a decrease in the mechanical properties and an increase in the swelling ratio of the hybrid. However, the swelling kinetics were accelerated due to a reduction in CS crystallinity. On the other hand, with increasing clay concentration, the increased cross-linked bonding mechanically reinforced the hybrid beyond the aforementioned adverse effect, to show improved tensile strength and a decrease in the swelling ratio. The voltage-induced deformation of hybrids became more pronounced with increasing applied voltage, but became less pronounced with increasing clay content under an applied electric field. After repeatedly switching the electric field on and off, the higher clay concentration (C(clay)>0.5wt.%) of the hybrid composites maintained the same capability of deswelling and swelling after more than 10 cycles, compared with both the pure CS film and the hybrid composites with lower clay content (e.g., 0.5wt.%). Compared with pure CS, a significant improvement in the anti-fatigue property against cyclic electric stimulations of the hybrid was found, which encourages the use of such a new class of hybrid composite in medical and pharmaceutical applications.

Local co-administration of gene-silencing RNA and drugs in cancer therapy: State-of-the art and therapeutic potential.

Gene-silencing miRNA and siRNA are emerging as attractive therapeutics with potential to suppress any genes, which could be especially useful in combination cancer therapy to overcome multidrug resistant (MDR) cancer. Nanomedicine aims to advance cancer treatment through functional nanocarriers that delivers one or more therapeutics to cancer tissue and cells with minimal off-target effects and suitable release kinetics and dosages. Although much effort has gone into developing circulating nanocarriers with targeting functionality for systemic administration, another alternative and straightforward approach is to utilize formulations to be administered directly to the site of action, such as pulmonary and intratumoral delivery. The combination of gene-silencing RNA with drugs in nanocarriers for localized delivery is emerging with promising results. In this review, the current progress and strategies for local co-administration of RNA and drug for synergistic effects and future potential in cancer treatment are presented and discussed. Key advances in RNA-drug anticancer synergy and localized delivery systems were combined with a review of the available literature on local co-administration of RNA and drug for cancer treatment. It is concluded that advanced delivery systems for local administration of gene-silencing RNA and drug hold potential in treatment of cancer, depending on indication. In particular, there are promising developments using pulmonary delivery and intratumoral delivery in murine models, but further research should be conducted on other local administration strategies, designs that achieve effective intracellular delivery and maximize synergy and feasibility for clinical use.

Inhibition of Periodontitis Induction Using a Stimuli-Responsive Hydrogel Carrying Naringin.

BACKGROUND: Developing a drug carrier with favorable handling characteristics that can respond to environmental changes after inflammation, such as pH changes, may be beneficial for treating periodontitis. This study aims to investigate the preclinical feasibility of using naringin, a naturally derived polymethoxylated flavonoid compound with anti-inflammatory properties, to inhibit periodontitis induction via a thermogelling and pH-responsive injectable hydrogel. METHODS: The hydrogel was made of amphipathic carboxymethyl-hexanoyl chitosan (CHC), ß-glycerol phosphate (ß-GP), and glycerol. Thermogelling and pH-responsive characteristics of the hydrogel, as well as cell viability after treatment with the hydrogel containing naringin, were evaluated in vitro. Hydrogel was subgingivally delivered when experimental periodontitis was induced in vivo, and therapeutic effect was evaluated with microcomputed tomography imaging, histology, and expression of inflammation-associated genes, including toll-like receptor (TLR)2, the receptor for advanced glycation end products (RAGE), myeloid differentiation primary response gene-88, and tumor necrosis factor (TNF)-α. RESULTS: The hydrogel was consistently fluidic at 4°C but rapidly gelled at 37°C. Release of naringin was faster at pH 5.5 to 6.5, and viability was significantly promoted by treatment with 0.85% naringin. Naringin-carrying CHC-ß-GP-glycerol hydrogel sites showed significantly reduced periodontal bone loss (P <0.05) and inflammatory infiltration (P <0.01) as well as significantly downregulated TLR2 (P <0.05), RAGE (P <0.01), and TNF-α (P <0.05) relative to the sites with experimental periodontitis alone. CONCLUSION: Naringin-carrying CHC-ß-GP-glycerol colloidal hydrogel can be used to inhibit induction of experimental periodontitis with favorable handling and inflammation-responsive characteristics.

Study on drug release behaviour of CDHA/chitosan nanocomposites--effect of CDHA nanoparticles.

To explore the effect of nanofiller-polymer interaction on the drug release behaviour from a monolithic membrane prepared by Ca-deficient hydroxyapatite (CDHA)/chitosan nanocomposite, release kinetics was investigated in terms of different synthetic processes, i.e. in situ and ex situ routes, and various amounts of CDHA. It was found that a higher value of diffusion exponent (n) was obtained for the membranes in situ synthesized compared with those ex situ prepared. In addition, the n value of the membranes in situ synthesized increased with increasing CDHA amount, which remained in the range below 10wt.%. However, as CDHA content exceeded 30%, the n value remained constant. It indicates that the drug diffusion mechanism is altered by the CDHA-chitosan interaction which is strongly influenced by both the synthesis process and the concentration of the CDHA nanofiller in the membrane. On the other hand, a lower permeability (P) value of the membranes was observed for those prepared via the in situ process. Furthermore, P value decreased and increased with increasing CDHA amount in the range below and above 10wt.%, respectively. It demonstrates that CDHA nanofillers act either diffusion barrier or diffusion enhancer for the CDHA/chitosan membranes, which is determined by the concentration of CDHA nanofiller and the synthesis route of nanocomposite.