Acid-conjugated Chitosan Nanoparticles Research Articles

Bile acid conjugated chitosan nanoparticles promote the proliferation and epithelial-mesenchymal transition of hepatocellular carcinoma by regulating the PI3K/Akt/mTOR pathway

Bile acids have been known to play significant roles at certain physiological levels in gastrointestinal metabolism. Yet, they are known to be carcinogenic and aid in tumor progression in most cases, although the roles remain uncertain. Hence, we tested the cytotoxic potential of cholic acid (CA) loaded chitosan nanoparticles (CNPs) on Hep3B cells. The physicochemical properties of the CNPs synthesized with CA load (CA-CNPs) were determined using standard techniques such as ultraviolet–visible spectrophotometry (UV–Vis), fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), dynamic light scattering (DLS) and transmission electron microscopy (TEM). The characteristic peak for chitosan nanoparticles were observed for plain CNPs (pCNPs) and CA-CNPs at around 300 nm as per UV–Vis analysis. FTIR analysis indicated the possible trapping of CA onto CNPs as certain peaks were retained and some peaks were shifted. XRD analysis determined that the peaks representing CA and pCNPs were collectively obtained in CA-CNPs. As per DLS analysis, the particle size, PDI and ζ-potential of the CA-CNPs were 259 nm, 0.284 and 30.4 mV. Further, the CA-CNPs were non-cytotoxic on Hep3B cells at the maximum tested concentration of 500 μg/mL. The viability at 500 μg/mL of CA-CNPs was two-fold higher than 500 μg/mL of pCNPs. Also, the pCNPs were not hemolytic and therefore could not have played a role in the increase of viability after treatment with CA-CNPs, which indicates that CA posed a major role in increased viability of Hep3B cells. As per quantitative PCR (qPCR), the upregulated gene expressions of PI3K, Akt, mTORC2, cMyc, Fibronectin, hVPS34, Slug and ZEB1 and the downregulated expression of the tumor suppressor PTEN indicates that PI3K/Akt/mTOR pathway mediated the induction of epithelial-to-mesenchymal transition (EMT) in response to CA-CNPs treatment on Hep3B cells.

Capecitabine Folic Acid Conjugated Chitosan Nanoparticles in CRC Targetting

Capecitabine Folic Acid Conjugated Chitosan Nanoparticles in CRC Targetting

In Vivo Cancer Microenvironment Responsive Glycan Receptor-Targeted Nanoparticles for Gemcitabine Delivery to Benzo[a]pyrene-Induced Lung Cancer Model.

Targeted gemcitabine (GEB) loaded 5-N-acetyl-neuraminic acid (Neu5Ac) assembled chitosan nanoparticles (CA-NPs) were formulated by ionotropic gelation process and evaluated for physicochemical and morphological characterization, in vitro and in vivo studies in A-549 cells and lung cancer mice model, respectively. The mean diameter of GEB-CA-Neu5Ac-NPs determined by dynamic light scattering was 161.16 ± 7.70 nm with a polydispersity index (PDI) value of 0.303 ± 0.011 and its zeta potential and entrapment efficiency (%EE) were 40.3 ± 3.45 mv and 66.11 ± 1.94%, respectively. The in vitro cellular uptake studies showed that glycan receptor-targeted nanoparticles deliver significantly more amount (p < 0.001) of GEB into the A-549 lung cancerous cells than non-targeted nanoparticles. The cytotoxicity study using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay clearly demonstrated that GEB-CA-Neu5Ac-NPs have lower IC50 value (6.39 ± 3.78 µg/ml) than others groups that showed that the greater lung cancerous cells inhibition potential of targeted nanoparticles. The in vivo biodistribution of the GEB-loaded 5-N-acetyl-neuraminic acid conjugated chitosan nanoparticles was revealed that targeted nanoparticles showed higher accumulation and retention for an extended period of time due to the active targeting ability of Neu5Ac to glycan receptors. Histopathological examination showed significant recovery in the physiological architecture upon administration of targeted nanoparticles. The glycan receptor-targeted nanoparticles treated groups showed a significant decline in the number of metastatic lung epithelial cells, as compared to the untreated positive control group (p < 0.001) confirming higher anticancer efficacy of the GEB-CA-Neu5Ac-NPs.

Dual targeting pH responsive chitosan nanoparticles for enhanced active cellular internalization of gemcitabine in non-small cell lung cancer

Lung cancer (LC), related with the enhanced expression of epidermal growth factor receptor (EGFR) and sialic acid binding receptors (glycan) brought about the development of EGFR and glycan receptor specific anticancer therapeutics. The current study assessed the formulation, physiochemical characterization, in vitro and in vivo effects of sialic acid (SA) and cetuximab (Cxmab) decorated chitosan nanoparticles (CSN-NPs) loaded with gemcitabine (GMC) targeted to glycan and EGFR over-expressing non-small-cell lung-cancer (NSCLC) A-549 cells. Chitosan (CSN) was conjugated with sialic acid via EDC/NHS chemistry followed by gemcitabine loaded sialic acid conjugated chitosan nanoparticles (GMC-CSN-SA-NPs) were prepared by ionic gelation method decorated with Cxmab by electrostatic interaction. In vitro cytotoxicity of NPs quantified using cell based MTT, DAPI and Annexing-V/PI apoptosis assays showed superior antiproliferative activity of targeted nanoformulations (GMC-CSN-SA-Cxmab-NPs ≫ GMC-CSN-SA-NPs, GMC-CSN-Cxmab-NPs) over non-targeted nanoformulation (GMC-CSN-NPs) against A-549 cells. In vivopharmacokinetic study showed superior bioavailability and in vivo therapeutic efficacy investigation exhibited strongest anticancer activity of glycan and EGFR targeted NPs (GMC-CSN-SA-Cxmab-NPs). GMC-CSN-SA-Cxmab-NPs demonstrated enhanced cellular internalization and better therapeutic potential, by specifically targeting glycan and EGFR on NSCLC A-549 cells and B[a]P induced lung cancer mice model, hence it might be a good substitute for non-targeted, conventional chemotherapy.

Sialic Acid Conjugated Chitosan Nanoparticles: Modulation to Target Tumour Cells and Therapeutic Opportunities.

Targeted delivery of therapeutics forestalls the dreadful delocalized effects, drug toxicities and needless immunosuppression. Cancer cells are bounteous with sialic acid and the differential expression of glycosyl transferase, glycosidase and monosaccharide transporter compared to healthy tissues. The current study entails the development and characterisation of sialic acid (SA)-labelled chitosan nanoparticles encapsulating gemcitabine (GEM). Chitosan (CS) was conjugated with SA using coupling reaction and characterised spectroscopically. Furthermore, different concentrations of chitosan and tripolyphosphate (TPP) were optimised to fabricate surface modified chitosan nanoparticles. SA conjugated chitosan nanoparticles encapsulating GEM (SA-CS_GEM NPs) of 232 ± 9.69nm with narrow distribution (PDI < 0.5) and zeta potential of - 19 ± 0.97mV was fabricated. GEM was successfully loaded in the SA-CS NPs, depicting prolonged and biphasic drug release pattern more elated at low pH. Pronounced cellular uptake (FITC tagged) and cytotoxicity (IC50 487.4nM) was observed in SA-CS_GEM NPs against A549 cells. IC50 for SA-CS_GEM NPs plunged with an increase in the time points from 24 to 72h. Concentration-dependent haemolytic study confirmed significant haemocompatibility of SA-CS_GEM NPs. Pharmacokinetic study was performed on Sprague-Dawley rats and the kinetic parameters were calculated using PKSolver 2.0. Results demonstrated a consequential refinement of 2.98 times in modified SA-CS_GEM NPs with a significant increase in retention time, bioavailability and elimination half-life, and decrease in elimination rate constant and volume of distribution in comparison to CS_GEM NPs. Therefore, SA-CS shell core nanoparticles could be a beneficial approach to target and treat NSCLC (non-small cell lung cancer) and direct for research possibilities to target the other tumour cells.

Kappa-Carrageenan Crosslinked Magnetic Folic Acid-Conjugated Chitosan Nanocomposites for Arginase Encapsulation, Delivery and Cancer Therapy

In this study, the Aspergillus niger L-arginase was expressed in Pichia pastorise and immobilized on Kappa-carrageenan crosslinked magnetic folic acid-conjugated chitosan nanoparticles (magnetic-FA/CTS NPs). L-arginase gene was isolated and cloned in pGAPZ[Formula: see text]A plasmid and transformed in P. pastoris. The purified L-arginase was loaded on magnetic-FA/CTS NPs nanocomposites and the structure of the synthesized nano-carriers was investigated using SEM, FT-IR and DLS techniques. Prepared formulations showed high encapsulation efficiency (91.93%). In vitro release and cytotoxic studies were performed at different pH (pH of 4–8) and against Molt-4 cancer cell line, respectively.

Phenylboronic acid-conjugated chitosan nanoparticles for high loading and efficient delivery of curcumin

In order to achieve high loading and efficient delivery of curcumin, phenylboronic acid-conjugated chitosan nanoparticles were prepared by a simple desolvation method. These nanoparticles exhibited a regular spherical shape with the average size about 200−230 nm and narrow size distribution, which were kinetically stable under physiological condition. Due to boronate ester formation between curcumin and phenylboronic acid groups in the nanoparticles, and the hydrogen bonding interactions between curcumin and nanocarriers, curcumin was successfully loaded into the nanoparticles with high drug loading content. These curcumin-loaded nanoparticles showed pH and reactive oxygen species (ROS)-triggered drug release behavior. In vitro cell experiments revealed that the blank nanoparticles were completely nontoxic to cultured cells, and the curcumin-loaded nanoparticles exhibited efficient antitumor efficiency against cancer cells. Moreover, the drug-loaded nanoparticles performed an enhanced growth inhibition in three-dimensional multicellular tumor spheroids. Thus, these nanocarriers would be a promising candidate for curcumin delivery in tumor treatment.

Amino acid conjugated chitosan nanoparticles for the brain targeting of a model dipeptidyl peptidase-4 inhibitor

Saxagliptin (SAX) is a dipeptidyl peptidase-4 enzyme inhibitor molecule now explored for its activity in the therapy of Alzheimer’s disease. Being extremely hydrophilic, it is unable to permeate the blood-brain barrier by the conventional therapy modalities. Further repurposing the drug, SAX is associated with a reduction in the blood sugar level in the periphery. In the present study, the chitosan-l-valine conjugate was synthesized by carbodiimide chemistry. The conjugate was then used to prepare nanoparticles encapsulating SAX. The nanoparticles were characterized by particle size, surface morphology, and entrapment efficiency. The stability of the formulations was determined by incubation with rat plasma and brain homogenate. The blood brain barrier permeability of the nanoparticles was successfully demonstrated by the incorporation of fluorescent dye, Rhodamine B in the nanoparticles. In vivo studies were conducted in rats and the results showed that the nanoparticles were highly stable in the plasma releasing only a minute amount of SAX (2.5 ng/mL) which was less than the Cmax of the pure SAX (51 ng/mL). The brain uptake studies showed an accumulation of 53 ng/mL SAX from the nanoparticles whereas the pure SAX showed no detectable amount of the drug after 24 h. The pharmacokinetic studies demonstrated that nanoparticles had an (AUC0–t) of 3.42 times lower than the pure SAX, indicating the stability of the prepared formulation in the plasma.

A multi-responsive biomimetic nano-complex platform for enhanced gene delivery.

RNA interference (RNAi) is widely regarded as a promising technology for disease treatment, yet one major obstacle for its clinical application is the lack of enhanced siRNA delivery vehicles to circumvent complex extra- and intracellular barriers. By integrating unique peculiarities of thioglycolic acid conjugated chitosan nanoparticles (TCS NPs), biomimetic transfersomes (T) and amphiphilic hyaluronic acid (HA-GMS), a novel nano-complex was prepared, where vascular endothelial growth factor (VEGF) siRNA loaded TCS NPs were cloaked by transfersomes with HA-GMS assembled on the surface (HT-TCS-siRNA NPs). The nano-complex provided superior siRNA protection and desirable stability at pH 7.4 and 6.5 (mimicking tumor tissue) and exerted proton sponge effects at acidic pH 5.0 (mimicking endo/lysosomes). The TCS NPs were stable at pH 5.0 but disintegrated in the presence of 10 mM glutathione (GSH) at pH 7.4 (mimicking tumor cytosol), which was favorable to release siRNA to the cytoplasm. In vitro cell uptake and gene silencing assays exhibited enhanced intracellular siRNA accumulation and VEGF silencing efficacy of HT-TCS-siRNA NPs in HeLa cells. The enhanced gene delivery capacity of the multi-responsive biomimetic nano-complex gives them potential for application in cancer therapy.

Development and characterization of folic acid-conjugated chitosan nanoparticles for targeted and controlled delivery of gemcitabinein lung cancer therapeutics

The present study was designed to investigate the tumor-targeting potential of gemcitabine (GEM)-loaded surface-tailored chitosan (CS)/poly (ethylene glycol) nanoparticles (FA-PEG-GEM-NPs). The nanoparticles encapsulated with GEM were prepared, characterized, and tethered with folic acid. The developed formulations were characterized with respect to particle size/poly-dispersity index, shape, and zeta potential analysis. The in vitro study shows the sustained drug-release kinetics during 48 h. The present result shows remarkable cytotoxicity rendered by GEM when delivered through FA-PEG-GEM-NPs formulation. The microscopic assessment is suggestive of significant uptake of FA-PEG-GEM-NPs in comparison with the unmodified PEG-GEM-NPs and free drug. Finally, our results advocate for the sizeable compatibility, comparatively less organ toxicity, and higher anti-tumor activity of ligand-anchored and PEGylated CS nanoparticles in vitro and corroborated by in vivo investigations. In conclusion, it is interpreted that surface-tailored nanoparticles are capable to ferry bioactives selectively and specifically to tumor sites with the interception of minimal side effects, thereby suggesting their potential application in cancer therapeutics.

Cross-Linked Dependency of Boronic Acid-Conjugated Chitosan Nanoparticles by Diols for Sustained Insulin Release.

Boronic acids have been widely investigated for their potential use as glucose sensors in glucose responsive polymeric insulin delivery systems. Interactions between cyclic diols and boronic acids, anchored to polymeric delivery systems, may result in swelling of the delivery system, releasing the drug. In this study, 4-formylphenylboronic acid conjugated chitosan was formulated into insulin containing nanoparticles via polyelectrolyte complexation. The nanoparticles had an average diameter of 140 ± 12.8 nm, polydispersity index of 0.17 ± 0.1, zeta potential of +19.1 ± 0.69 mV, encapsulation efficiency of 81% ± 1.2%, and an insulin loading capacity of 46% ± 1.8% w/w. Changes in size of the nanoparticles and release of insulin were type of sugar- and concentration-dependent. High concentration of diols resulted in a sustained release of insulin due to crosslink formation with boronic acid moieties within the nanoparticles. The formulation has potential to be developed into a self-regulated insulin delivery system for the treatment of diabetes.

Phenylboronic Acid-Mediated Tumor Targeting of Chitosan Nanoparticles

The phenylboronic acid-conjugated chitosan nanoparticles were prepared by particle surface modification. The size, zeta potential and morphology of the nanoparticles were characterized by dynamic light scattering, zeta potential measurement and transmission electron microscopy. The cellular uptake, tumor penetration, biodistribution and antitumor activity of the nanoparticles were evaluated by using monolayer cell model, 3-D multicellular spheroid model and H22 tumor-bearing mice. The incorporation of phenylboronic acid group into chitosan nanoparticles impart a surface charge-reversible characteristic to the nanoparticles. In vitro evaluation using 2-D and 3-D cell models showed that phenylboronic acid-decorated nanoparticles were more easily internalized by tumor cells compared to non-decorated chitosan nanoparticles, and could deliver more drug into tumor cells due to the active targeting effect of boronic acid group. Furthermore, the phenylboronic acid-decorated nanoparticles displayed a deeper penetration and persistent accumulation in the multicellular spheroids, resulting in better inhibition growth to multicellular spheroids than non-decorated nanoparticles. Tumor penetration, drug distribution and near infrared fluorescence imaging revealed that phenylboronic acid-decorated nanoparticles could penetrate deeper and accumulate more in tumor area than non-decorated ones. In vivo antitumor examination demonstrated that the phenylboronic acid-decorated nanoparticles have superior efficacy in restricting tumor growth and prolonging the survival time of tumor-bearing mice than free drug and drug-loaded chitosan nanoparticles.

Molecular Mechanism of Enhanced Anticancer Effect of Nanoparticle Formulated LY2835219 via p16-CDK4/6-pRb Pathway in Colorectal Carcinoma Cell Line

LY2835219 is a dual inhibitor to CDK4 and CDK6. This study was to prepare LY2835219-loaded chitosan nanoparticles (CNP/LY) and LY2835219-loaded hyaluronic acid-conjugated chitosan nanoparticles (HACNP/LY) and revealed their anticancer effect and influence on p16-CDK4/6-pRb pathway against colon cell line. The nanoparticle sizes of CNP/LY and HACNP/LY were approximately195±39.6 nm and217±31.1 nm, respectively. The zeta potentials of CNP/LY and HACNP/LY were37.3±1.5 mV and30.3±2.2 mV, respectively. And the preparation process showed considerable drug encapsulation efficiency and loading efficiency. LY2835219, CNP/LY, and HACNP/LY inhibited HT29 cell proliferation with 0.68, 0.54, and 0.30 μM of IC50, respectively. G1 phase was arrested by LY2835219 and its formulations. Furthermore, inhibition of CDK4/6 by LY2835219 formulations induced CDK4, CDK6, cyclin D1, and pRb decrease and p16 increase at both protein and mRNA levels. Overall, nanoparticle formulated LY2835219 could enhance the cytotoxicity and cell cycle arrest, and HACNP/LY strengthened the trend furtherly compared to CNP/LY. It is the first time to demonstrate the anticancer effect and mechanism against HT29 by LY2835219 and its nanoparticles. The drug and its nanoparticle formulations delay the cell growth and arrest cell cycle through p16-CDK4/6-pRb pathway, while the nanoparticle formulated LY2835219 could strengthen the process.

Folic Acid Conjugated Chitosan Nanoparticles for Tumor Targeting of Therapeutic and Imaging Agents

Anticancer drugs are typically distributed non-specifically in the body, where they affect both cancerous and normal cells. This limits the drug level achievable within the tumor, compromising the therapeutic efficacy, and results in potential toxic effects on normal tissues. Targeted delivery of chemotherapeutics exclusively to cancer cells is the focus of intensive research for improvement of anticancer therapy. Various drug delivery systems have been investigated for this purpose, with therapeutic-carrying polymeric nanoparticulate systems designed for specific targeting of tumor cells receiving special interest. Chitosan, a natural polymer derived from crustacean shells, has attracted particular attention as a drug carrier. The simple and mild preparation methods, low toxicity, good stability, controlled drug release and the ability to overcome biological barriers have made chitosan-based nanoparticles popular in drug and gene delivery applications. Chitosan nanoparticles have been fabricated with optimal size and surface characteristics in order to tailor the behavior within the biological system, including circulation time, as well as passive and active cancer targeting. Folic acid is widely employed as a ligand targeting cancerous cells as its receptor which ‘shuttles’ folic acid into the cells via endocytosis is over-expressed on the surface of many human epithelial cancer cells. Incorporating folic acid into chitosan-based drug and gene delivery formulations renders the systems with an efficient targeting capacity. Furthermore, it is possible to formulate chitosan nanocarriers that display multiple useful characteristics extending beyond targeted delivery. The versatility of these systems is also being exploited in nanotheranostics. Keywords: Chitosan, diagnostic and therapeutic imaging, drug and gene delivery, folic acid.

Erratum to: Preparation, characterization, and in vitro release of folic acid-conjugated chitosan nanoparticles loaded with methotrexate for targeted delivery

To reduce the toxicity of methotrexate (MTX) and increase the targeting of nanoparticles, the MTX-loaded chitosan (CS) covalently bonded with folic acid (FA) nanoparticles were prepared, and sodium tripolyphosphate was used as the cross-linking agent. FA was successfully conjugated to CS confirmed by 1H-NMR and Fourier transform infrared spectrometer (FT-IR). The prepared FA–CS nanoparticles were characterized by FT-IR spectroscopy to confirm the cross-linking reaction between FA–CS and cross-linking agent. X-ray diffraction was performed to reveal the crystalline nature of the drug after encapsulation. The average diameters of the nanoparticles ranged from 293.9 ± 24.2 to 401.5 ± 20.4 nm with a narrow particle size distribution. In vitro release pattern in phosphate buffer saline (pH 6.8) indicated that the characteristics of the MTX-loaded nanoparticles appeared to have an initial burst effect and followed by a slow, sustained drug release. FA or low molecular weight FA conjugate fragments were also released from the nanoparticles, which might have potential to reduce toxic effects of MTX within the body.

Preparation, characterization, and in vitro release of folic acid-conjugated chitosan nanoparticles loaded with methotrexate for targeted delivery

Preparation, characterization, and in vitro release of folic acid-conjugated chitosan nanoparticles loaded with methotrexate for targeted delivery

Alginate-folic acid-modified chitosan nanoparticles for photodynamic detection of intestinal neoplasms

Colorectal cancer is one of the leading causes of cancer death and often goes undetected with current colonoscopy practices. Improved methods of detecting dysplasia and tumors during colonoscopy could significantly improve mortality. Herein, we report a high-performance nanoparticle for photodynamic detection of colorectal cancer, where alginate is physically complexed with folic acid-modified chitosan to form nanoparticles with improved drug release in the cellular lysosome. The incorporated alginate molecules could complex stably with chitosan via electrostatic attraction, and the z-average diameter and zeta-potential of the prepared nanoparticles (fCAN) was 115 nm and 22 mV, respectively, enough to keep the nanoparticles stable in aqueous suspension without aggregation. When loaded with 5-aminolevulinic acid (5-ALA; 27% loading efficiency), the nanoparticles (fCANA) displayed no differences in particle size or zeta-potential compared to fCAN. Moreover, the fCANA nanoparticles were readily taken up by colorectal cancer cells via folate receptor-mediated endocytosis. Subsequently, the loaded 5-ALA was release in the lysosome, and this was promoted by the reduced attraction intensity between chitosan and 5-ALA via the deprotonated alginate, resulting in a higher intracellular PpIX accumulation for the photodynamic detection. These studies demonstrate that the alginate incorporated and folic acid-conjugated chitosan nanoparticles are excellent vectors for colorectal-specific delivery of 5-ALA for fluorescent endoscopic detection.

Folic Acid-Conjugated Chitosan Nanoparticles Enhanced Protoporphyrin IX Accumulation in Colorectal Cancer Cells

Folic acid can be covalently conjugated to chitosan molecules via its gamma-carboxyl moiety and thus retain a high affinity for colorectal cancer cells bearing folate receptor overexpression. Colorectal cancer is one of the leading causes of malignant death and often goes undetected with current colonoscopy practices. Improved methods of detecting dysplasia and tumors during colonoscopy will improve mortality. A folic acid conjugated chitosan nanoparticle as a suitable vehicle for carrying 5-aminolaevulinic acid (5-ALA) is developed to enhance the detection of colorectal cancer cells in vivo after a short-term uptake period. Chitosan can be successfully conjugated with folic acid to produce folic acid-chitosan conjugate, which is then loaded with 5-ALA to create nanoparticles (fCNA). The loading efficiency of 5-ALA in fCNA particles and the z-average diameter were in the range 35-40% and 100 nm, respectively. The zeta-potential for fCNA was 20 mV, enough to keep the nanoparticle stable without aggregation. The fCNA is then incubated with HT29 and Caco-2 colorectal cancer cell lines overexpressing folate receptor on the surface of the cell membrane to determine the rate of accumulation of protoporphyrin IX (PpIX). The results show that fCNA can be taken up more easily by HT29 and Caco-2 cell lines after short-term uptake period, most likely via receptor-mediated endocytosis, and the PpIX accumulates in cancer cells as a function of the folate receptor expression and the folic acid modification. Therefore, the folic acid-chitosan conjugate appears to be an ideal vector for colorectal-specific delivery of 5-ALA for fluorescent endoscopic detection.