Tailoring Chlorthalidone Aqueous Solubility by Cocrystallization: Stability and Dissolution Behavior of a Novel Chlorthalidone-Caffeine Cocrystal.

A cocrystal of the antihypertensive drug chlorthalidone (CTD) with caffeine (CAF) was obtained (CTD-CAF) by the slurry method, for which a 2:1 stoichiometric ratio was found by powder and single-crystal X-ray diffraction analysis. Cocrystal CTD-CAF showed a supramolecular organization in which CAF molecules are embedded in channels of a 3D network of CTD molecules. The advantage of the cocrystal in comparison to CTD is reflected in a threefold solubility increase and in the dose/solubility ratios, which diminished from near-unit values for D0D to 0.29 for D0CC. Furthermore, dissolution experiments under non-sink conditions showed improved performance of CTD-CAF compared with pure CTD. Subsequent studies showed that CTD-CAF cocrystals transform to CTD form I where CTD precipitation inhibition could be achieved in the presence of pre-dissolved polymer HPMC 80-120 cPs, maintaining supersaturation drug concentrations for at least 180 min. Finally, dissolution experiments under sink conditions unveiled that the CTD-CAF cocrystal induced, in pH-independent manner, faster and more complete CTD dissolution when compared to commercial tablets of CTD. Due to the stability and dissolution behavior of the novel CTD-CAF cocrystal, it could be used to develop solid dosage forms using a lower CTD dose to obtain the same therapeutic response and fewer adverse effects.

Nanoconfinement of a Pharmaceutical Cocrystal with Praziquantel in Mesoporous Silica: The Influence of the Solid Form on Dissolution Enhancement.

Nanoconfinement is a recent strategy to enhance solubility and dissolution of active pharmaceutical ingredients (APIs) with poor biopharmaceutical properties. In this work, we combine the advantage of cocrystals of racemic praziquantel (PZQ) containing a water-soluble coformer (i.e., increased solubility and supersaturation) and its confinement in a mesoporous silica material (i.e., increased dissolution rate). Among various potential cocrystalline phases of PZQ with dicarboxylic acid coformers, the cocrystal with glutaric acid (PZQ-GLU) was selected and successfully loaded by the melting method into nanopores of SBA-15 (experimental pore size of 5.6 nm) as suggested by physical and spectroscopic characterization using various complementary techniques like N2 adsorption, powder X-ray diffraction (PXRD), infrared spectroscopy (IR), solid-state NMR (ss-NMR), differential scanning calorimetry (DSC), and field emission-scanning electron microscopy (FE-SEM) analysis. The PZQ-GLU phase confined in SBA-15 presents more mobility according to ss-NMR studies but still retains its cocrystal-like features in the IR spectra, and it also shows depression of the melting transition temperature in DSC. On the contrary, pristine PZQ loaded into SBA-15 was found only in the amorphous state, according to the aforementioned studies. This dissimilar behavior of the composites was attributed to the larger crystal lattice of PZQ over the PZQ-GLU cocrystal (3320.1 vs 1167.9 Å3) and to stronger intermolecular interactions between PZQ and GLU, facilitating the confinement of a more mobile solid-like phase in the constrained channels. Powder dissolution studies under extremely nonsink conditions (SI = 0.014) of the confined PZQ-GLU and amorphous PZQ phases embedded in mesoporous silica showed transient supersaturation behavior when dissolving in simulated gastric fluid (HCl pH 1.2 at 37 ± 0.5 °C) in a similar fashion to the bare cocrystal PZQ-GLU. A comparison of the area under the curve (AUC0-90 min) of the dissolution profiles afforded a dissolution advantage of 2-fold (p < 0.05) of the new solid phases over pristine racemic PZQ after 90 min; under these conditions, the solubilized API reprecipitated as the recently discovered PZQ hemihydrate (PZQ-HH). In the presence of a cellulosic polymer, sustained solubilization of PZQ from composites SBA-15/PZQ or SBA-15/PZQ-GLU was observed, increasing AUC0-90 min up to 5.1-fold in comparison to pristine PZQ. The combination of a confined solid phase in mesoporous silica and a methylcellulose polymer in the dissolution medium effectively maintained the drug solubilized during times significant to promote absorption. Finally, powder dissolution studies under intermediate nonsink conditions (SI = 1.99) showed a fast release profile from the nanoconfined PZQ-GLU phase in SBA-15, which reached rapid saturation (95% drug dissolved at 30 min); the amorphous PZQ composite and bare PZQ-GLU also displayed an immediate release of the API but at a lower rate (69% drug dissolved at 30 min). In all of these cases, a large dissolution advantage was observed from any of the novel solid phases over PZQ.

Dissolution Advantage of Nitazoxanide Cocrystals in the Presence of Cellulosic Polymers.

The effect of hydroxypropyl methylcellulose (HPMC) and methylcellulose (Methocel® 60 HG) on the dissolution behavior of two cocrystals derived from nitazoxanide (NTZ), viz., nitazoxanide-glutaric acid (NTZ-GLU, 1:1) and nitazoxanide-succinic acid (NTZ-SUC, 2:1), was explored. Powder dissolution experiments under non-sink conditions showed similar dissolution profiles for the cocrystals and pure NTZ. However, pre-dissolved cellulosic polymer in the phosphate dissolution medium (pH 7.5) modified the dissolution profile of NTZ when starting from the cocrystals, achieving transient drug supersaturation. Subsequent dissolution studies under sink conditions of polymer-based pharmaceutical powder formulations with NTZ-SUC cocrystals gave a significant improvement of the apparent solubility of NTZ when compared with analogous formulations of pure NTZ and the physical mixture of NTZ and SUC. Scanning electron microscopy and powder X-ray diffraction analysis of samples recovered after the powder dissolution studies showed that the cocrystals undergo fast dissolution, drug supersaturation and precipitation both in the absence and presence of polymer, suggesting that the solubilization enhancement is due to polymer-induced delay of nucleation and crystal growth of the less soluble NTZ form. The study demonstrates that the incorporation of an appropriate excipient in adequate concentration can be a key factor for inducing and maintaining the solubilization of poorly soluble drugs starting from co-crystallized solid forms. In such a way, cocrystals can be suitable for the development of solid dosage forms with improved bioavailability and efficacy in the treatment of important parasitic and viral diseases, among others.

Interrelation of the dissolution behavior and solid-state features of acetazolamide cocrystals.

The thermal behavior, phase stability, indicative stability and intrinsic dissolution rates of a series of cocrystals and cocrystal hydrates derived from the pharmaceutically active ingredient acetazolamide (ACZ) and 2-aminobenzamide (2ABAM), 2,3-dihydroxybenzoic acid (23DHBA), 2-hydroxybenzamide (2HBAM), 4-hydroxybenzoic acid (4HBA), nicotinamide (NAM) and picolinamide (PAM) as cocrystal formers have been evaluated. Upon heating in an inert atmosphere most of the cocrystals tested demonstrated first the elimination of the crystal former, followed by ACZ degradation. Only in cocrystals with NAM was melting observed. Under controlled temperature and relative humidity conditions all cocrystals tested were stable. However, phase stability tests in a medium simulating physiological conditions (HCl 0.01N, pH2.0) indicated that cocrystals ACZ-NAM-H2O and ACZ-PAM gradually transform into ACZ. All cocrystals examined gave enhanced intrinsic dissolution rates when compared to pure ACZ and the largest dissolution rate constants were measured for the cocrystals that transformed in the phase stability test (approximate two-fold increase of the dissolution rate constants). The series of cocrystals examined herein exhibits an inverse correlation between the intrinsic dissolution rates and the melting/decomposition temperatures as well as the dimension of the hydrogen-bonded ACZ aggregates found in the corresponding crystal structure, indicating that solid-state stability is the major influence on dissolution performance.

Sulfonate salts of the therapeutic agent dapsone: 4-[(4-aminophenyl)sulfonyl]anilinium benzenesulfonate monohydrate and 4-[(4-aminophenyl)sulfonyl]anilinium methanesulfonate monohydrate.

Dapsone, formerly used to treat leprosy, now has wider therapeutic applications. As is the case for many therapeutic agents, low aqueous solubility and high toxicity are the main problems associated with its use. Derivatization of its amino groups has been widely explored but shows no significant therapeutic improvements. Cocrystals have been prepared to understand not only its structural properties, but also its solubility and dissolution rate. Few salts of dapsone have been described. The title salts, C12H13N2O2S(+)·C6H5O3S(-)·H2O and C12H13N2O2S(+)·CH3SO3(-)·H2O, crystallize as hydrates and both compounds exhibit the same space group (monoclinic, P21/n). The asymmetric unit of each salt consists of a 4-[(4-aminophenyl)sulfonyl]anilinium monocation, the corresponding sulfonate anion and a water molecule. The cation, anion and water molecule form hydrogen-bonded networks through N-H...O=S, N-H...Owater and Owater-H...O=S hydrogen bonds. For both salts, the water molecules interact with one sulfonate anion and two anilinium cations. The benzenesulfonate salt forms a two-dimensional network, while the hydrogen bonding within the methanesulfonate salt results in a three-dimensional network.

Enhanced cellular uptake and gene silencing activity of siRNA molecules mediated by chitosan-derivative nanocomplexes.

The RNA interference (RNAi) constitutes a conservative mechanism in eukaryotic cells that induces silencing of target genes. In mammalians, the RNAi is triggered by siRNA (small interfering RNA) molecules. Due to its potential in silencing specific genes, the siRNA has been considered a potential alternative for the treatment of genetic and acquired diseases. However, the siRNA therapy has been limited by its low stability and rapid degradation in presence of nucleases, low cellular uptake, and immune response activation. In order to overcome these drawbacks, we propose the synthesis and characterization of non-viral delivery systems using chitosan derivatives to obtain siRNA complexes (polyplexes). The non-viral delivery systems synthesized included PEG-g-OCs (oligochitosan) and PEG-g-Cs (chitosan medium molecular weight). Both systems allowed the formation of siRNA polyplexes, increased the stability of siRNA in the presence of nucleases, enhanced cellular internalization, and showed low toxicity in the A549 cell line. Finally, the complexes obtained with the PEG-g-OCs system showed silencing activity in a GFP model in the cell line A549 in comparison with naked siRNA.

Design and in vitro evaluation of a new nano-microparticulate system for enhanced aqueous-phase solubility of curcumin.

Curcumin, a yellow polyphenol derived from the turmeric Curcuma longa, has been associated with a diverse therapeutic potential including anti-inflammatory, antioxidant, antiviral, and anticancer properties. However, the poor aqueous solubility and low bioavailability of curcumin have limited its potential when administrated orally. In this study, curcumin was encapsulated in a series of novel nano-microparticulate systems developed to improve its aqueous solubility and stability. The nano-microparticulate systems are based entirely on biocompatible, biodegradable, and edible polymers including chitosan, alginate, and carrageenan. The particles were synthesized via ionotropic gelation. Encapsulating the curcumin into the hydrogel nanoparticles yielded a homogenous curcumin dispersion in aqueous solution compared to the free form of curcumin. Also, the in vitro release profile showed up to 95% release of curcumin from the developed nano-microparticulate systems after 9 hours in PBS at pH 7.4 when freeze-dried particles were used.

Formulation approaches to short interfering RNA and MicroRNA: challenges and implications.

RNA interference has emerged as a potentially powerful tool in the treatment of genetic and acquired diseases by delivering short interfering RNA (siRNA) or microRNA (miRNA) to target genes, resulting in their silencing. However, many physicochemical and biological barriers have to be overcome to obtain efficient in vivo delivery of siRNA and miRNA molecules to the organ/tissue of interest, thereby enabling their effective clinical therapy. This review discusses the challenges associated with the use of siRNA and miRNA and describes the nonviral delivery strategies used in overcoming these barriers. More specifically, emphasis has been placed on those technologies that have progressed to clinical trials for both local and systemic siRNA and miRNA delivery.

A Novel Aerosol Method for the Production of Hydrogel Particles.

A novel method of generating hydrogel particles for various applications including drug delivery purposes was developed. This method is based on the production of hydrogel particles from sprayed polymeric nano/microdroplets obtained by a nebulization process that is immediately followed by gelation in a crosslinking fluid. In this study, particle synthesis parameters such as type of nebulizer, type of crosslinker, air pressure, and polymer concentration were investigated for their impact on the mean particle size, swelling behavior, and morphology of the developed particles. Spherical alginate-based hydrogel particles with a mean particle size in the range from 842 to 886 nm were obtained. Using statistical analysis of the factorial design of experiment it was found that the main factors influencing the size and swelling values of the particles are the alginate concentration and the air pressure. Thus, it was demonstrated that the method described in the current study is promising for the generation of hydrogel particles and it constitutes a relatively simple and low-cost system.

Risks and benefits of commonly used herbal medicines in Mexico.

In Mexico, local empirical knowledge about medicinal properties of plants is the basis for their use as home remedies. It is generally accepted by many people in Mexico and elsewhere in the world that beneficial medicinal effects can be obtained by ingesting plant products. In this review, we focus on the potential pharmacologic bases for herbal plant efficacy, but we also raise concerns about the safety of these agents, which have not been fully assessed. Although numerous randomized clinical trials of herbal medicines have been published and systematic reviews and meta-analyses of these studies are available, generalizations about the efficacy and safety of herbal medicines are clearly not possible. Recent publications have also highlighted the unintended consequences of herbal product use, including morbidity and mortality. It has been found that many phytochemicals have pharmacokinetic or pharmacodynamic interactions with drugs. The present review is limited to some herbal medicines that are native or cultivated in Mexico and that have significant use. We discuss the cultural uses, phytochemistry, pharmacological, and toxicological properties of the following plant species: nopal (Opuntia ficus), peppermint (Mentha piperita), chaparral (Larrea divaricata), dandlion (Taraxacum officinale), mullein (Verbascum densiflorum), chamomile (Matricaria recutita), nettle or stinging nettle (Urtica dioica), passionflower (Passiflora incarnata), linden flower (Tilia europea), and aloe (Aloe vera). We conclude that our knowledge of the therapeutic benefits and risks of some herbal medicines used in Mexico is still limited and efforts to elucidate them should be intensified.

The functional evaluation of human peptide/histidine transporter 1 (hPHT1) in transiently transfected COS-7 cells.

Recently, the expression of the human peptide/histidine transporter (hPHT1, SLC15A4) mRNA was observed in the GI tract and in Caco-2 cells, suggesting that it may participate in the intestinal absorption of peptide-based agents. This study aims to elucidate the: (i) protein expression pattern of hPHT1 (SLC15A4) in human small intestine; (ii) cloning of the hPHT1 full-length sequence; (iii) functional characterization of hPHT1 in transiently transfected COS-7 cells. The expression of hPHT1 was measured using Western blot and immunohistochemical analysis. The hPHT1 full-sequence was amplified from BeWo cells, inserted into the pcDNA3.1-V5/His TOPO plasmid and transiently transfected into COS-7 cells to investigate the uptake kinetics of [3H]histidine and [3H]carnosine. Time, pH and sodium-dependent uptake studies were performed in mock (empty vector) and hPHT1-COS-7 cells. Results demonstrated hPHT1 protein expression in different intestinal regions. Histidine and carnosine uptake was linear in hPHT1-COS-7 cells over 15 min and was found to be pH-dependent. These substrates and valacyclovir showed significantly higher uptake at pH 5.0 in the hPHT1 transients when contrasted to the mock COS-7 cells, whereas glycylsarcosine uptake was significantly lower and unaffected by pH. Other di- and tripeptides also showed affinity for hPHT1. This study presents the initial functional characterization, the protein expression of the hPHT1 transporter and provides insight into a potentially different route for increasing peptide and peptide-based drug transport.

Delineation of human peptide transporter 1 (hPepT1)-mediated uptake and transport of substrates with varying transporter affinities utilizing stably transfected hPepT1/Madin-Darby canine kidney clones and Caco-2 cells.

In the present investigation, the uptake and transport kinetics of valacyclovir (VACV), 5-aminolevulinic acid (5-ALA), and benzylpenicillin (BENZ) were studied in stably transfected Madin-Darby canine kidney (MDCK)/human peptide transporter 1 (hPepT1)-V5&His clonal cell lines expressing varying levels of epitope-tagged hPepT1 protein (low, medium, and high expression) and in Caco-2 cells to delineate hPepT1-mediated transport kinetics. These compounds were selected due to the fact that they are known PepT1 substrates, yet also have affinity for other transporters. Caco-2 cells, traditionally used for studying peptide-based drug transport, were included for comparison purposes. The time, pH, sodium, and concentration dependence of cellular uptake and permeability were measured using mock, clonal hPepT1-MDCK, and Caco-2 cells. A pH-dependent effect was observed in the hPepT1-expressing clones and Caco-2 cells, with an increase of 1.96-, 1.84-, and 2.05-fold for VACV, 5-ALA, and BENZ uptake, respectively, at pH 6 versus 7.4 in the high-expressing hPepT1 cells. BENZ uptake was significantly decreased in Caco-2 and MDCK cells in Na(+)-depleted buffer, whereas VACV uptake only decreased in Caco-2 cells. Concentration-dependent uptake studies in the mock-corrected hPepT1-MDCK and Caco-2 cells demonstrated hPepT1 affinity ranking of VACV > 5-ALA > BENZ. The apical-to-basal apparent permeability coefficient (P(app)) values of VACV, 5-ALA, and BENZ in mock-corrected hPepT1-MDCK cells showed solely hPepT1-mediated transport in contrast to Caco-2 cells. Lower K(m) values and higher P(app) in Caco-2 cells compared with hPepT1-MDCK cells suggested the involvement of multiple transporters in Caco-2 cells. Thus, hPepT1-MDCK cells corrected for endogenous transporter expression may be a more appropriate model for screening compounds for their affinity to hPepT1.

A novel hPepT1 stably transfected cell line: establishing a correlation between expression and function.

Stably transfected MDCK/hPepT1-V5&His clonal cell lines expressing varying levels of epitope-tagged hPepT1 protein were established to quantify the relationship between transgene hPepT1 expression levels and its functional kinetics in facilitating peptide and peptide-like drug uptake and transport in vitro. The hPepT1 sequence was amplified from Caco-2 cell mRNA, inserted into the pcDNA3.1 -V5&His TOPO plasmid, and transfected into MDCK cells. Transgene protein levels were quantified by Western Blot analysis utilizing a standard curve generated with a positive control protein containing a V5&His epitope. Three clones expressing different levels of the hPepT1 fusion protein (low, medium, and high) were selected for the functional characterization with [14C]Gly-Sar and [3H]carnosine. The MDCK/hPepT1 cells expressed a novel hPepT1/epitope tag protein with an apparent molecular mass of 110 kDa. The [14C]Gly-Sar uptake in the transfected cells was sodium-independent and pH-dependent, demonstrating enhanced uptake, the rate of which increased significantly from the weakly to strongly expressing hPepT1 MDCK/hPepT1 -V5&His clones as compared to the mock cell line at pH 6.0. The uptake and permeability of [14C]Gly-Sar and [3H]carnosine demonstrated a direct correlation between the hPepT1 level of expression, uptake, and transport capabilities. Molecular and functional characterization of the MDCK/hPepT1-V5&His cell line confirmed a directly proportional relationship between Vmax and Papp versus the molar levels of hPepT1 transgene expression. This stably transfected hPepT1 cell line may serve as a useful in vitro model for screening and quantifying peptide and peptide-like drug transport as a function of hPepT1 expression in drug discovery.

Expression of multiple drug resistance conferring proteins in normal Chinese and Caucasian small and large intestinal tissue samples.

Multidrug resistance conferring proteins (MDRCP) are ATP-binding cassette (ABC) transporters known to significantly influence the absorption, distribution, metabolism, and elimination (ADME) and toxic behavior of many therapeutic agents. Research in the pharmacogenomics area has suggested that mutations and variable expression patterns of these MDCRPs may exist in tissue samples from different ethnic groups. The goal of this study was to examine the expression of P-glycoprotein (PGP), sister of PGP (S-PGP), multidrug resistance protein 3 (Mdr3), multidrug resistance like proteins 1-5 (MRP 1-5), and lung resistance associated protein (LRP) in tissue slides and protein lysates derived from normal adult small or large intestines of Caucasian or Chinese origin. Our results demonstrated ubiquitous expression of PGP, MRP 1, MRP 4, and LRP in the small and large intestinal epitheliums originating from both Caucasian and Chinese origin. S-PGP, Mdr3, MRP 2, and MRP 3 exhibited variable expression in the tissue slides and protein lysates derived from the Chinese and Caucasian small and large intestines. MRP 5 was not observed in any of the samples studied. The results suggest that MDCRPs may have distinct expression profiles in the small and large intestines that potentially vary with genetic background. These studies provide a foundation for further investigations to verify these findings across a wider number of patients of different ethnic backgrounds.

Current perspectives on established and putative mammalian oligopeptide transporters.

Peptides and peptide-based drugs are increasingly being utilized as therapeutic agents for the treatment of numerous disorders. The increasing development of peptide-based therapeutic agents is largely due to technological advances including the advent of combinatorial peptide libraries, peptide synthesis strategies, and peptidomimetic design. Peptides and peptide-based agents have a broad range of potential clinical applications in the treatment of many disorders including AIDS, hypertension, and cancer. Peptides are generally hydrophilic and often exhibit poor passive transcellular diffusion across biological barriers. Insights into strategies for increasing their intestinal absorption have been derived from the numerous studies demonstrating that the absorption of protein digestion products occurs primarily in the form of small di- and tripeptides. The characterization of the pathways of intestinal, transepithelial transport of peptides and peptide-based drugs have demonstrated that a significant degree of absorption occurs through the role of proteins within the proton-coupled, oligopeptide transporter (POT) family. Considerable focus has been traditionally placed on Peptide Transporter 1 (PepT1) as the main mammalian POT member regulating intestinal peptide absorption. Recently, several new POT members, including Peptide/Histidine Transporter 1 (PHT1) and Peptide/Histidine Transporter 2 (PHT2) and their splice variants have been identified. This has led to an increased need for new experimental methods enabling better characterization of the biophysical and biochemical barriers and the role of these POT isoforms in mediating peptide-based drug transport.