Ghanashyam Acharya

Hydrotropic Solubilization of Paclitaxel: Analysis of Chemical Structures for Hydrotropic Property

AbstractPurpose. To identify hydrotropic agents that can increase aqueous paclitaxel (PTX) solubility and to study the chemical structures necessary for hydrotropic properties so that polymeric hydrotropic agents can be synthesized. Methods. More than 60 candidate hydrotropic agents (or hydro- tropes) were tested for their ability to increase the aqueous PTX solubility. A number of nicotinamide analogues were synthesized based on the observation that nicotinamide showed a favorable hydrotropic property. The identified hydrotropes for PTX were used to examine the structure-activity relationship.Results. N,N-Diethylnicotinamide (NNDENA) was found to be the most effective hydrotropic agent for PTX. The aqueous PTX solubility was 39 mg/ml and 512 mg/ml at NNDENA concentrations of 3.5 M and 5.95 M, respectively. These values are 5-6 orders of magnitude greater than the intrinsic solubility of 0.30 ± 0.02 μg/ml. N-Picolylnicotinamide, N-allylnicotinamide, and sodium salicylate were also excellent hydrotropes for PTX. Solubility data showed that an effective hydrotropic agent should be highly water soluble while maintaining a hydrophobic segment. Conclusions. The present study identified several hydrotropic agents effective for increasing aqueous solubility of PTX and analyzed the structural requirements for this hydrotropic property. This information can be used to find other hydrotropic compounds and to synthesize polymeric hydrotropes that are effective for PTX and other poorly water-soluble drugs.

Hydrotropic polymer micelles containing acrylic acid moieties for oral delivery of paclitaxel

Hydrotropic polymers (HPs) and their micelles have been recently developed as vehicles for delivery of poorly water-soluble drugs, such as paclitaxel (PTX), by oral administration. The release of PTX from HP micelles, however, was slow and it took more than a day for complete release of the loaded PTX. Since the gastrointestinal (GI) transit time is known to be only several hours, pH-sensitive HP micelles were prepared for fast release of the loaded PTX responding to pH changes along the GI tract. Acrylic acid (AA) was introduced, as a release modulator, into HPs by copolymerization with 4-(2-vinylbenzyloxy)-N,N-(diethylnicotinamide) (VBODENA). The AA content was varied from 0% to 50% (in the molar ratio to VBODENA). HPs spontaneously produced micelles in water, and their critical micelle concentrations (CMCs) ranged from 31 microg/mL to 86 microg/mL. Fluorescence probe study using pyrene showed that blank HP micelles possessed a good pH sensitivity, which was clearly observed at relatively high AA contents and pH>6. The pH sensitivity also affected the PTX loading property. Above pH 5, the PTX loading content and loading efficiency in HP micelles were significantly reduced. Although this may be primarily due to the AA moieties, other factors may include PTX degradation and polymer aggregation. The PTX release from HP micelles with more than 20% (mol) AA contents was completed within 12 h in a simulated intestinal fluid (SIF, pH=6.5). The HP micelles without any AA moiety showed very slow release profiles. In the simulated gastric fluid (SGF, pH=1.6), severe degradation of the released PTX was observed. The pH-dependent release of PTX from HP micelles can be used to increase the bioavailability of PTX upon oral delivery.

The hydrogel template method for fabrication of homogeneous nano/microparticles

Nano/microparticles have been used widely in drug delivery applications. The majority of the particles are prepared by the conventional emulsion methods, which tend to result in particles with heterogeneous size distribution with sub-optimal drug loading and release properties. Recently, microfabrication methods have been used to make nano/microparticles with a monodisperse size distribution. The existing methods utilize solid templates for making particles, and the collection of individual particles after preparation has not been easy. The new hydrogel template approach was developed to make the particle preparation process simple and fast. The hydrogel template approach is based on the unique properties of physical gels that can undergo sol-gel phase transition upon changes in environmental conditions. The phase reversible hydrogels, however, are in general mechanically too weak to be treated as a solid material. It was unexpectedly found that gelatin hydrogels could be made to possess various properties necessary for microfabrication of nano/microparticles in large quantities. The size of the particles can be adjusted from 200 nm to >50 microm, providing flexibility in controlling the size in drug delivery formulations. The simplicity in processing makes the hydrogel template method useful for scale-up manufacturing of particles. The drug loading capacity is 50% or higher, and yet the initial burst release is minimal. The hydrogel template approach presents a new strategy of preparing nano/microparticles of predefined size and shape with homogeneous size distribution for drug delivery applications.

A study of drug release from homogeneous PLGA microstructures.

The hydrogel template method was used to fabricate homogeneous drug-PLGA microparticles. Four drugs (felodipine, risperidone, progesterone, and paclitaxel) were loaded into the PLGA particles with the homogeneous size of 10microm, 20microm, and 50microm. The drug loading into the PLGA microparticles was 50% and higher. The felodipine-PLGA microstructures of four different sizes showed that the drug release kinetics is dependent on the total surface area available for drug release. The smaller the particle size, the release rate was faster. Two types of microparticles (10microm diameter and 10microm height, and 50microm diameter and 5microm height) showed zero-order release and complete release was observed within 2weeks. The release rate, however, was not exactly proportional to the surface area. Different drugs which were loaded into the same PLGA formulation showed different release profiles. The main difference was on the initial burst release. The overall release profile seems to be similar for different drugs, if the release profile is adjusted to eliminate the burst release. The initial burst release appears to be inversely related to the water-solubility of a drug, i.e., the lower the water-solubility of a drug, the higher the burst release. The hydrogel template method allowed preparation of homogeneous particles with predefined sizes with high drug loading. It allowed study on the effect of size and shape on the drug release kinetics. With the microparticles of homogeneous size and shape, the drug release kinetics can be projected based on the size of microparticles and water-solubility of a drug. The ability of making homogeneous particles is expected to provide better prediction and reproducibility of the drug release property of a given formulation.

Detection of pathogenic E. coli O157:H7 by a hybrid microfluidic SPR and molecular imaging cytometry device.

Current methods to screen for bacterial contamination involve using costly reagents such as antibodies or PCR reagents or time‐costly growth in cultures. There is need for portable, real‐time, multiplex pathogen detection technology that can predict the safety of food. Surface plasmon resonance (SPR) imaging is a sensitive, label‐free method that can detect the binding of an analyte to a surface by the changes in refractive index that occur upon binding. We have designed a hybrid microfluidic biochip to perform multiplexed detection of single‐celled pathogens using a combination of SPR and fluorescence imaging. The device consists of an array of gold spots, each functionalized with a capture biomolecule targeting a specific pathogen. This biosensor array is enclosed by a polydimethylsiloxane microfluidic flow chamber that delivers a magnetically concentrated sample to be tested. The sample is imaged by SPR on the bottom of the biochip and epi‐fluorescence on the top. The prototype instrument was successfully able to image antibody‐captured E. coli O157:H7 bacteria by SPR and fluorescence imaging. The efficiency of capture of these bacteria by the magnetic particles was determined using spectrophotometric ferric oxide absorbance measurements. The binding of the E. coli to each spot was quantified by measuring the percent of the gold spot area upon which the bacteria was bound and analyzed using NIH ImageJ software. This hybrid imaging approach of pathogenic E. coli detection coupled with an estimate of relative infectivity is shown to be a working example of a testing device for potential foodborne pathogens.

Ocular drug delivery nanowafer with enhanced therapeutic efficacy.

Presently, eye injuries are treated by topical eye drop therapy. Because of the ocular surface barriers, topical eye drops must be applied several times in a day, causing side effects such as glaucoma, cataract, and poor patient compliance. This article presents the development of a nanowafer drug delivery system in which the polymer and the drug work synergistically to elicit an enhanced therapeutic efficacy with negligible adverse immune responses. The nanowafer is a small transparent circular disc that contains arrays of drug-loaded nanoreservoirs. The slow drug release from the nanowafer increases the drug residence time on the ocular surface and its subsequent absorption into the surrounding ocular tissue. At the end of the stipulated period of drug release, the nanowafer will dissolve and fade away. The in vivo efficacy of the axitinib-loaded nanowafer was demonstrated in treating corneal neovascularization (CNV) in a murine ocular burn model. The laser scanning confocal imaging and RT-PCR study revealed that once a day administered axitinib nanowafer was therapeutically twice as effective, compared to axitinib delivered twice a day by topical eye drop therapy. The axitinib nanowafer is nontoxic and did not affect the wound healing and epithelial recovery of the ocular burn induced corneas. These results confirmed that drug release from the axitinib nanowafer is more effective in inhibiting CNV compared to the topical eye drop treatment even at a lower dosing frequency.

In situ assembled diffraction grating for biomolecular detection

The authors report experiments with a diffraction-based biosensor based on self-assembly of target-containing nanobeads that form optical diffraction gratings. They demonstrate that the diffraction signal is a function of the bead size, and that noise is minimized by normalizing the intensities of the diffraction modes. They characterize the dependence of the diffraction signal on equivalent bead size and demonstrate the potential of the scheme in detecting biologically significant molecules.

Rapid Detection of S‐Adenosyl Homocysteine Using Self‐Assembled Optical Diffraction Gratings

Quantitative characterization of biomolecules is critical for molecular diagnostics and drug development. Several assays based on spectrophotometry, fluorometry, chemiluminescence, and electrochemical immunoassays have been reported for biomolecular detection. These methods are often slow owing to multiple sample pretreatment steps that increase analysis time and cost. Immunoassays that combine high sensitivity with fast, robust, and inexpensive methods for biomolecular detection are of growing importance. Herein the development of a self-assembled optical diffraction biosensor is described which is devoid of microfabrication or enzymatic amplification for the rapid detection of S-adenosyl homocysteine (SAH), a potential diagnostic marker for cardiovascular disease, with a sensitivity limit of 24.5 pgmL . SAH is a low-molecular-mass analyte (384 Da) consisting of the nucleoside adenine joined to the amino acid homocysteine (Hcy) by a 5’ thioether linkage. Our method relies on a sandwich-binding approach, wherein SAH is bound by an antibody through the Hcy moiety while an adenine-specific RNA aptamer binds to the adenine moiety (Figure 1). We designed a strategy using antibody-coupled (Ab) beads to capture SAH from solution in addition to aptamerfunctionalized micropatterns specific for adenine which are stamped on a gold-coated glass slide (gold chip). Based on these specific interactions, SAH bound to the Ab beads produces a self-assembled optical diffraction grating upon exposure to the aptamer-functionalized micropatterns (Figure 2). The high-affinity adenosine-specific aptamer containing the 39-mer sequence CGG AUG AGA CGC UUG GCG UGU GCU GUG GAG AGU CAU CCG was chosen

Dexamethasone nanowafer as an effective therapy for dry eye disease

Dry eye disease is a major public health problem that affects millions of people worldwide. It is presently treated with artificial tear and anti-inflammatory eye drops that are generally administered several times a day and may have limited therapeutic efficacy. To improve convenience and efficacy, a dexamethasone (Dex) loaded nanowafer (Dex-NW) has been developed that can release the drug on the ocular surface for a longer duration of time than drops, during which it slowly dissolves. The Dex-NW was fabricated using carboxymethyl cellulose polymer and contains arrays of 500 nm square drug reservoirs filled with Dex. The in vivo efficacy of the Dex-NW was evaluated using an experimental mouse dry eye model. These studies demonstrated that once a day Dex-NW treatment on alternate days during a five-day treatment period was able to restore a healthy ocular surface and corneal barrier function with comparable efficacy to twice a day topically applied dexamethasone eye drop treatment. The Dex-NW was also very effective in down regulating expression of inflammatory cytokines (TNF-α, and IFN-γ), chemokines (CXCL-10 and CCL-5), and MMP-3, that are stimulated by dry eye. Despite less frequent dosing, the Dex-NW has comparable therapeutic efficacy to topically applied Dex eye drops in experimental mouse dry eye model, and these results provide a strong rationale for translation to human clinical trials for dry eye.

Hydrogel templates for the fabrication of homogeneous polymer microparticles.

Nano/microparticulate drug delivery systems with homogeneous size distribution and predefined shape are important in understanding the influence of the geometry and dimensions of these systems on blood circulation times and cellular uptake. We present a general method using water dissolvable hydrogel templates for the fabrication of homogeneous, shape-specific polymer/drug constructs in the size range of 200 nm to 50 μm. This hydrogel template strategy is mild, inexpensive, and readily scalable for the fabrication of multifunctional drug delivery vehicles.