Non-Invasive Intranasal Delivery of pApoE2: Effect of Multiple Dosing on the ApoE2 Expression in Mice Brain.

Chitosan-based polymeric micelles are promising non-viral nanocarriers for safe and targeted gene delivery. Multi-functionalized chitosan polymeric micelles were prepared by grafting fatty acid, cell-penetrating peptide, and mannose on the chitosan backbone. The polymeric micelles were subjected to surface morphology and surface topography using scanning electron microscopy and atomic force microscopy, respectively. The hemotoxic profile of the prepared polymeric micelles was established against erythrocytes and was found to be <5% hemotoxic up to the concentration of 600 µg/mL. In vitro ApoE2 expression in primary astrocytes and neurons was analyzed. Multi-functionalized polymeric micelles produced greater (p < 0.05) transfection in astrocytes and neurons in comparison to mono-functionalized micelles. Intranasal administration of polymeric micelles/pApoE2 polyplex led to significantly higher (p < 0.05) in vivo pApoE2 expression than chitosan and unfunctionalized polymeric micelles-treated mice groups. The outcomes of this study predict that the developed multi-functionalized polymeric micelles could be an effective and safe gene delivery platform to the brain through the intranasal route.

Multifunctionalized Cationic Chitosan Polymeric Micelles Polyplexed with pVGF for Noninvasive Delivery to the Mouse Brain through the Intranasal Route for Developing Therapeutics for Alzheimer's Disease.

Multifunctionalized Chitosan-based polymeric micelles were used to deliver pVGF to the brain. VGF (non-acronymic) plays significant roles in neurogenesis and learning as well as synaptic and cognitive functions. Therefore, VGF gene therapy could be a better approach in developing effective therapeutics against Alzheimer's disease. Multifunctionalized chitosan polymeric micelles were developed by grafting oleic acid (OA) on the chitosan (CS) skeleton followed by penetratin (PEN) and mannose (MAN) conjugation. The OA-g-CS-PEN-MAN graft polymer formed cationic nanomicelles in an aqueous medium and polyplexed with pVGF. The polymeric micelles were nontoxic and cationic in charge and had an average hydrodynamic diameter of 199.8 ± 15.73 nm. Qualitative in vitro transfection efficiency of OA-g-CS-PEN-MAN/pGFP polyplex was investigated in bEnd.3, primary neurons, and astrocyte cells. In vivo transfection efficiency of OA-g-CS-PEN-MAN/pVGF polyplexes was analyzed in C57BL6/J mice after intranasal administration for 7 days. The VGF expression levels in primary astrocytes and neurons after OA-g-CS-PEN-MAN/pVGF treatment were 2.4 ± 0.24 and 1.49 ± 0.02 pg/µg of protein, respectively. The VGF expression in the OA-g-CS-PEN-MAN/pVGF polyplex-treated animal group was 64.9 ± 12.7 pg/mg of protein, significantly higher (p < 0.01) than that of the unmodified polymeric micelles. The in vivo transfection outcomes revealed that the developed multifunctionalized OA-g-CS-PEN-MAN polymeric micelles could effectively deliver pVGF to the brain, transfect brain cells, and express VGF in the brain after noninvasive intranasal administration.

Synthesis and Characterization of Fatty Acid Grafted Chitosan Polymeric Micelles for Improved Gene Delivery of VGF to the Brain through Intranasal Route.

Multifunctional fatty acid grafted polymeric micelles are an effective and promising approach for drug and gene delivery to the brain. An alternative approach to bypass the blood-brain barrier is administration through intranasal route. Multifunctional fatty acid grafted polymeric micelles were prepared and characterized for pVGF delivery to the brain. In vitro pVGF expression was analyzed in bEnd.3 cells, primary astrocytes, and neurons. Comparative in-vivo pVGF expression was analyzed to evaluate the effective route of administration between intranasal and intravenous. Biocompatible, multifunctional polymeric micelles were prepared, having an average size of 200 nm, and cationic zeta potential. Modified polymers were found to be hemo- and cyto-compatible. When transfected with the different modified chitosan formulations, significantly (p < 0.05) higher VGF expression was observed in primary astrocytes and neurons using the mannose, Tat peptide, and oleic acid grafted chitosan polymer. Compared to intravenous administration, intranasal administration of pVGF in polyplex formulation led to significantly (p < 0.05) higher pVGF expression. Developed multifunctional polymeric micelles were an effective pVGF delivery platform to the brain. Mannose and Tat ligand tagging improved the pVGF delivery to the brain.

PEGylated Dendrimer Mediated Delivery of Bortezomib: Drug Conjugation versus Encapsulation.

Poor aqueous solubility of anticancer drug bortezomib (BTZ) still remains a major challenge in the development of a successful formulation. The dendrimeric platform can provide a better opportunity to deliver BTZ with improved solubility. BTZ encapsulated in PEGylated PAMAM dendrimers (BTZ-PEG-PAMAM) was characterized and evaluated comparatively with encapsulated and conjugated dendritic formulations. The particle size of BTZ-PEG-PAMAM was 188.6 ± 4.17 nm, with entrapment efficiency of 78.61 ± 2.91% and drug loading of 39.30 ± 1.98%. The aqueous solubility of BTZ in PAMAM-PEG conjugate was enhanced by 68.11 folds in comparison to pure drug. In vitro drug release profile was found to be sustained up to 72 h. A comparative colorimetric MTT assay against A549 and MCF-7 cancer cells resulted in maximum efficacy from BTZ-PEG-PAMAM with IC50 value 333.14 ± 15.42 and 152.60 ± 24.56 nM, respectively. Significantly higher cellular internalization was observed in FITC tagged BTZ-PEG-PAMAM. In vivo pharmacokinetic study performed on Sprague Dawley rats resulted in 8.63 folds increase in bioavailability for BTZ-PEG-PAMAM than pure drug. Pharmacokinetic parameters of BTZ-PEG-PAMAM were better and improved over BTZ and other dendritic formulations. In conclusion, the prepared formulation of BTZ-PEG-PAMAM has given significant outcome and this strategy may be further explored for better delivery of BTZ in future.

Lactoferrin Coupled Lower Generation PAMAM Dendrimers for Brain Targeted Delivery of Memantine in Aluminum-Chloride-Induced Alzheimer's Disease in Mice.

Lower generation PAMAM dendrimers have an immense potential for drug delivery with lower toxicity, but these dendrimers yet need certain basic ameliorations. In this study, the brain delivery potential of the synthesized PAMAM-Lf (lower generation PAMAM and lactoferrin conjugate) loaded with memantine (MEM) was explored and evaluated in vitro and in vivo in the disease-induced mouse model. The developed nanoscaffolds were characterized for size, zeta potential and in vitro release. Increase in the average size from 11.54 ± 0.91 to 131.72 ± 4.73 nm, respectively, was observed for drug-loaded PAMAM (i.e., PAMAM-MEM) and PAMAM-Lf (i.e., MEM-PAMAM-Lf).  Release profile of MEM from MEM-PAMAM-Lf was slow and sustained up to 48 h. In vivo biodistribution in the Sprague-Dawley rat model revealed that the brain uptake of MEM-PAMAM-Lf was significantly higher than that of MEM alone. The behavioral response study in the healthy rats did not result in any significant changes. The in vivo study in an AlCl3-induced Alzheimer's (AD) mice model showed a significant improvement in behavioral responses. Optical density, which reflects the acetylcholinesterase (AChE) activity, was highest in the AL group 0.16 ± 0.01 (higher than the CON group, 0.09 ± 0.02; p < 0.05). No significant suppression of AChE activity was recorded in all the other treated groups. Similarly, the DOPAmine and 3,4 dihydroxyphenylacetic acid (DOPAC) levels were unaffected by the developed formulations. The study reported improved brain bioavailability of MEM in AlCl3-induced Alzheimer's mice leading to improved memory, with the resultant mechanism behind in a descriptive manner. This study is among the preliminary studies reporting the memory improvement aspect of PAMAM-Lf conjugates for MEM in AlCl3-AD induced mice. The formulation developed was beneficial in AD-induced mice and had a significant impact on the memory aspects.

Correction to: Radiolabeled PLGA Nanoparticles for Effective Targeting of Bendamustine in Tumor Bearing Mice.

The typesetter did not use the Fig. 6 provided by the author with his proof corrections, and instead duplicated Fig. 7 by the Fig. 6 caption. The original article has been corrected.

Radiolabeled PLGA Nanoparticles for Effective Targeting of Bendamustine in Tumor Bearing Mice.

PURPOSE: Bendamustine is an important drug for the treatment of chronic lymphatic leukaemia (CLL), non-Hodgkin lymphoma (NHL). However, its delivery is challenging due to its instability. Current approach reports the development and characterization of bendamustine encapsulated PLGA nanoparticles for the effective targeting to leukemic cells. METHODS: The prepared, bendamustine loaded PLGA nanoparticles (BLPNP) were developed and characterized for particle size, zeta potential and polydispersity index. The formed nanoparticles were further characterized with the help of electron microscopy for surface morphology. The formed nanoparticles were evaluated for cytotoxicity, cell uptake, ROS and cell apoptosis against THP-1 leukemic cells as a part of in vitro evaluation. In vivo organ bio-distribution and tumor regression studies were performed to track in vivo behaviour of BLPNP. RESULTS: The average particle size was 138.52 ± 3.25 nm, with 0.192 ± 0.036 PDI and - 25.4 ± 1.38 mV zeta potential. TEM images revealed the homogeneous particle size distribution with uniform shape. In vitro release exhibited a sustained drug-release behaviour up to 24 h. Cytotoxicity against THP-1 cells through MTT assay observed IC50 value of 27.8 ± 2.1 µM for BLPNP compared to pure drug, which was 50.42 ± 3.4 µM. Moreover, in vitro studies like cell-uptake and cell apoptosis studies further confirmed the higher accumulation of BLPNP in comparison to the pure drug. Organ distribution and tumor regression studies were performed to track in vivo behaviour of bendamustine loaded nanoparticles. CONCLUSION: The overall study described a promising approach in terms of safety, least erythrocytic toxicity, better IC50 value with enhance tumor targeting and regression.

Bendamustine-PAMAM Conjugates for Improved Apoptosis, Efficacy, and in Vivo Pharmacokinetics: A Sustainable Delivery Tactic.

Successful delivery of a chemotherapeutic agent like bendamustine still remains a challenge in clinical conditions like chronic lymphatic leukemia (CLL), non-Hodgkin lymphoma (NHL), and multiple myeloma. We have conjugated bendamustine to polyamidoamine (PAMAM) dendrimers after conjugating with N-(hydroxyethyl)maleimide (spacer) via an ester bond. The particle size of PAMAM-bendamustine conjugate was 49.8 ± 2.5 nm. In vitro drug release resulted in sustained release with improved solution stability of drug up to 72 h. In a 24 h cytotoxicity study by MTT assay against human monoblastic leukemia cells (THP-1), the IC50 value for PAMAM-bendamustine was 32.1 ± 4.8 µM compared to 50.42 ± 3.4 µM and 2303 ± 106.5 µM for bendamustine and PAMAM dendrimer, respectively. Significantly higher cell uptake and apoptosis were observed in THP-1 cells by PAMAM-bendamustine conjugate which was confirmed by flow cytometry and confocal laser scanning microscopy. Preliminary in vivo studies undertaken included pharmacokinetics studies, organ distribution studies, and tumor inhibition studies. In healthy Wistar rat model (1CBM IV push model), the pharmacokinetic studies revealed that bioavailability and t1/2 increased significantly, i.e., almost 8.5-fold (193.8 ± 1.116 vs 22.8 ± 0.158 µg mL-1/h) and 5.1-fold (0.75 ± 0.005 vs 3.85 ± 0.015 h), respectively, for PAMAM-bendamustine conjugate compared to pure bendamustine ( p < 0.05), however, clearance and volume of distribution were found to be decreased compared to those of free drug. The study suggests that PAMAM-bendamustine conjugate was not only stable for the longer period but also least toxic and highly taken up by THP-1 cells to exert an anticancer effect at the reduced dose. Tumor inhibition and biodistribution studies in tumor-bearing BALB/c mice revealed that PAMAM-bendamustine conjugate was more effective than the pure drug and showed higher accumulation in the tumor.

Polymeric Nanocarriers: A New Horizon for the Effective Management of Breast Cancer.

BACKGROUND: Delivery of chemotherapeutic drugs for the diagnosis and treatment of cancer is becoming advanced day by day. However, the challenge of the effective delivery system still does exist. In various types of cancers, breast cancer is the most commonly diagnosed cancer among women. Breast cancer is a combination of different diseases. It cannot be considered as only one entity because there are many specific patient factors, which are involved in the development of this disease. Nanotechnology has opened a new area in the effective treatment of breast cancer due to the several benefits offered by this technology. METHODS: Polymeric nanocarriers are among one of the effective delivery systems, which has given promising results in the treatment of breast cancers. Nanocarriers does exert their anticancer effect either through active or passive targeting mode. RESULTS: The use of nanocarriers has been resolute about the adverse effects of chemotherapeutic drugs such as poor solubility and less penetrability in tumor cells. CONCLUSION: The present review is focused on recent developments regarding polymeric nanocarriers, such as polymeric micelles, polymeric nanoparticles, dendrimers, liposomes, nanoshells, fullerenes, carbon nanotubes (CNT) and quantum dots, etc. for their recent advancements in breast cancer therapy.

Dendrimer encapsulated and conjugated delivery of berberine: A novel approach mitigating toxicity and improving in vivo pharmacokinetics.

Berberine (BBR) is a nitrogenous cyclic natural alkaloid with potential anticancer activity. However it has been less explored due to its poor pharmacokinetic profile. Dendrimers (e.g. PAMAM) have promising potential to deliver anticancer drugs/bio-actives because of their well-defined architecture, monodispersity and tailor-made surface functionality. In the present study it was attempted to deliver berberine through G4 PAMAM dendrimers by conjugation (BPC) as well as encapsulation (BPE) approach. The developed encapsulated and conjugated berberine formulations were found to have size in the approximate range of 100-200nm while zeta potential was almost same as PAMAM G4 dendrimer. The entrapment efficiency in BPE was found to be 29.9%, whereas, the percentage conjugation in BPC was found to be 37.49% indicating high drug payload in conjugation. The developed nano-formulations were characterized through 1H NMR, FT-IR as well as electron microscopy (SEM and TEM). The in vitro release study in different media (water and PBS 7.4) showed sustained release pattern of BBR. Almost 72% and 98% drug was released within 24h respectively; whereas in PBS almost 80% and 98% release was observed within 24h, respectively. The formulations followed Higuchi release and first order release as best fit release kinetic model. MTT assay results showed significantly higher anticancer activity for the PAMAM-BBR (BPC) (p<0.01) against MCF-7 and MDA-MB-468 breast cancer cells. The time dependent ex vivo hemolytic toxicity of the BPC and BPE was significantly less (<5%) even after 24h, which indicated that the formulations can be regarded as significantly safe and biocompatible. Similarly, the in vivo hematological parameters were analyzed through auto-analyzer and the formulations were found to be safer and biocompatible with very least but insignificant (p>0.05) effects. The in vivo pharmacokinetic parameters were found to be impressively improved in albino rat model. The pharmacokinetic parameters such as half-life (t1/2) and AUC of berberine were impressively improved in the plasma level time in vivo studies in albino rat model. The obtained t1/2 was 14.33h for BPC compared to 6.7h for BBR alone. The overall conclusion says that among both the developed formulations the conjugated formulation (BPC) was found to be more prominent than the encapsulated one (BPE). Therefore conclusively conjugation can be a better option for the delivery of natural bio-actives through dendrimers.

Enhanced apoptotic and anticancer potential of paclitaxel loaded biodegradable nanoparticles based on chitosan.

Taxanes have established and proven effectivity against different types of cancers; in particular breast cancers. However, the high hemolytic toxicity and hydrophobic nature of paclitaxel and docetaxel have always posed challenges to achieve safe and effective delivery. Use of bio-degradable materials with an added advantage of nanotechnology could possibly improve the condition so as to achieve better and safe delivery. In the present study paclitaxel loaded chitosan nanoparticles were formulated and optimized using simple w/o nanoemulsion technique. The observed average size, pdi, zeta potential, entrapment efficiency and drug loading for the optimized paclitaxel loaded chitosan nanoparticle formulation (PTX-CS-NP-10) was 226.7±0.70nm, 0.345±0.039, 37.4±0.77mV, 79.24±2.95% and 11.57±0.81%; respectively. Nanoparticles were characterized further for size by Transmission Electron Microscopy (TEM). In vitro release studies exhibited sustained release pattern and more than 60% release was observed within 24h. Enhanced in vitro anticancer activity was observed as a result of MTT assay against triple negative MDA-MB-231 breast cancer cell lines. The observed IC50 values obtained for PTX-CS-NP-10 was 9.36±1.13µM and was almost 1.6 folds (p<0.05) less than the pure drug. Similarly, PTX-CS-NP-10 were extremely biocompatible and safe as observed for haemolytic toxicity which was almost 4 folds less (p<0.05) than the naïve drug. Anticancer activity was further evaluated using flow cytometry for apoptosis. Cell apoptosis study revealed that PTX-CS-NP-10 treatment resulted into enhanced (almost double) late cell apoptosis than naïve paclitaxel. Hence the developed nanoparticulate formulation not only reduced the overall toxicity but also resulted into improved anticancer efficacy of paclitaxel. It can be concluded that a robust, stable and comparatively safe nanoformulation of paclitaxel was developed, characterized and evaluated.

Polypropyleneimine and polyamidoamine dendrimer mediated enhanced solubilization of bortezomib: Comparison and evaluation of mechanistic aspects by thermodynamics and molecular simulations.

Bortezomib (BTZ) is the first proteasome inhibitor approved by the US-FDA is majorly used for the treatment of newly diagnosed and relapsed multiple myeloma including mantle cell lymphoma. BTZ is hydrophobic in nature and is a major cause for its minimal presence as marketed formulations. The present study reports the design, development and characterization of dendrimer based formulation for the improved solubility and effectivity of bortezomib. The study also equally focuses on the mechanistic elucidation of solubilization by two types of dendrimers i.e. fourth generation of poly (amidoamine) dendrimers (G4-PAMAM-NH2) and fifth generation of poly (propylene) imine dendrimers (G5-PPI-NH2). It was observed that aqueous solubility of BTZ was concentration and pH dependent. At 2mM G5-PPI-NH2 concentration, the fold increase in bortezomib solubility was 1152.63 times in water, while approximately 3426.69 folds increase in solubility was observed at pH10.0, respectively (p<0.05). The solubility of the drug was increased to a greater extent with G5-PPI-NH2 dendrimers because it has more hydrophobic interior than G4-PAMAM-NH2 dendrimers. The release of BTZ from G5-PPI-NH2 complex was comparatively slower than G4-PAMAM-NH2. The thermodynamic treatment of data proved that dendrimer drug complexes were stable at all pH with values of ΔG always negative. The experimental findings were also proven by molecular simulation studies and by calculating RMSD and intermolecular hydrogen bonding through Schrodinger software. It was concluded that PPI dendrimers were able to solubilize the drug more effectively than PAMAM dendrimers through electrostatic interactions.

Dendrimer nanoarchitectures for cancer diagnosis and anticancer drug delivery.

Dendrimers are novel nanoarchitectures with unique properties including a globular 3D shape, a monodispersed unimicellar nature and a nanometric size range. The availability of multiple peripheral functional groups and tunable surface engineering enable the facile modification of the dendrimer surface with different therapeutic drugs, diagnostic agents and targeting ligands. Drug encapsulation, and solubilizing and passive targeting also equally contribute to the therapeutic use of dendrimers. In this review, we highlight recent advances in the delivery of anticancer drugs using dendrimers, as well as other biomedical and diagnostic applications. Taken together, the immense potential and utility of dendrimers are envisaged to have a significant positive impact on the growing arena of drug delivery and targeting.

PLGA Nanoparticles and Their Versatile Role in Anticancer Drug Delivery.

Nanotechnological advancement has become a key standard for the diagnosis and treatment of several complex disorders such as cancer by utilizing the enhanced permeability and retention effect and tumor-specific targeting. Synthesis and designing the formulation of active agents in terms of their efficient delivery is of prime importance for healthcare. The use of nanocarriers has resolved the undesirable characteristics of anticancer drugs such as low solubility and poor permeability in cells. Several types of nanoparticles (NPs) have been designed with the use of various polymers along or devoid of surface engineering for targeting tumor cells. All NPs include polymers in their framework and, of these, polylactide-co-glycolide (PLGA) is biodegradable and Food and Drug Administration approved for human use. PLGA has been used extensively in the development of NPs for anticancer drug delivery. The extensive use of PLGA NPs is promising for cancer therapy, with higher efficiency and less adverse effects. The present review focused on recent developments regarding PLGA NPs, the methods used for their preparation, their characterization, and their utility in the delivery of chemotherapeutic agents.

Biodegradable nano-architectural PEGylated approach for the improved stability and anticancer efficacy of bendamustine.

Bendamustine is a drug of choice for the treatment of several cancers including non- Hodgkin lymphoma (NHL) and Chronic Lymphocytic Leukemia (CLL). The unstable nature of the drug, however, offers a major obstacle in its effective formulation development. The present study was aimed to achieve improved stability and efficacy of bendamustine via co-polymeric PEG-PLGA nanoparticulate approach. PEG-PLGA co-polymeric conjugate was synthesized and characterized by FT-IR and 1H NMR spectroscopy. Bendamustine loaded nanoparticles (PLGA and PEG-PLGA) were prepared, optimized and characterized for size, zeta and electron microscopy (SEM and TEM). The average size, pdi (polydispersity index), zeta potential and entrapment efficiency for bendamustine loaded PEG-PLGA nanoparticles (PPBNP 15) was 297.3±2.055nm, 0.256±0.012, -6.62±0.081mV and 52.30±3.66%, respectively. The in vitro release studies displayed sustained release nature of bendamustine. The Krosmeyer-Peppas model was the best fit model as a result of kinetic modelling for in vitro release. The ex vivo hemolytic toxicity of the PPBNP 15 was significantly less (approx. 11%; 4 folds) compared to pure drug (p<0.05). The cytotoxicity study showed significantly higher anticancer activity against MCF-7, T47D and PC-3 cells (p<0.05) compared to naïve bendamustine. The developed biodegradable nanoparticles improved the stability of bendamustine and were equally stable, less toxic and highly effective against different cancerous cells.

Polymeric Micelles: Recent Advancements in the Delivery of Anticancer Drugs.

Nanotechnology, in health and medicine, extensively improves the safety and efficacy of different therapeutic agents, particularly the aspects related to drug delivery and targeting. Among various nano-carriers, polymer based macromolecular approaches have resulted in improved drug delivery for the diseases like cancers, diabetes, autoimmune disorders and many more. Polymeric micelles consisting of hydrophilic exterior and hydrophobic core have established a record of anticancer drug delivery from the laboratory to commercial reality. The nanometric size, tailor made functionality, multiple choices of polymeric micelle synthesis and stability are the unique properties, which have attracted scientists and researchers around the world to work upon in this opportunistic drug carrier. The capability of polymeric micelles as nano-carriers are nowhere less significant than nanoparticles, liposomes and other nanocarriers, as per as the commercial feasibility and presence is concerned. In fact polymeric micelles are among the most extensively studied delivery platforms for the effective treatment of different cancers as well as non-cancerous disorders. The present review highlights the sequential and recent developments in the design, synthesis, characterization and evaluation of polymeric micelles to achieve the effective anticancer drug delivery. The future possibilities and clinical outcome have also been discussed, briefly.