Development of rectal enema as microbicide (DREAM): Preclinical progressive selection of a tenofovir prodrug enema.

HIV pre-exposure prophylaxis (PrEP) strategies have the potential to prevent millions of incident HIV infections each year. However, the efficacy of PrEP strategies has been plagued by issues of non-adherence, likely because of the difficulty in motivating otherwise healthy people to adhere to treatment regimens that require significant behavioral changes and daily discipline. An alternative approach to PrEP is to focus on strategies that fit in to normal, and even desirable, sexual behaviors, such as the use of cleansing enemas by men who have sex with men (MSM) prior to receptive anal intercourse (RAI). Here, we describe preclinical efforts toward optimizing a tenofovir (TFV)-based enema formulation for rectal PrEP. Using a murine model, we compared the plasma and tissue pharmacokinetics of TFV and various TFV prodrugs, including tenofovir disoproxil fumarate (TDF), tenofovir alafenamide (TAF), and hexadecyloxypropyl tenofovir (CMX157), after dosing as enema formulations with varying osmolality and ion content. We observed that the enema vehicle composition played a more important role than the TFV prodrug properties in achieving rapid and therapeutically relevant tenofovir diphosphate (TFV-DP) concentrations in mouse colorectal tissue. Our results support the next steps, which are further preclinical (non-human primate) and clinical development of a hypo-osmolar TFV enema product for rectal PrEP.

Mucus-penetrating budesonide nanosuspension enema for local treatment of inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a chronic inflammatory gastrointestinal disorder that affects more than 1 million individuals in the USA. Local therapy with enema formulations, such as micronized budesonide (Entocort®), is a common strategy for treating patients with distally active IBD. However, we hypothesize that micronized particulates are too large to effectively penetrate colorectal mucus, limiting the extent of drug delivery to affected tissues prior to clearance. Here, we describe the development of a budesonide nanosuspension (NS) with the appropriate surface coating and size to enhance penetration of colorectal mucus and ulcerated colorectal tissues. We demonstrate that model fluorescent polystyrene (PS) particles ∼200 nm in size with a muco-inert Pluronic F127 coating provide enhanced mucosal distribution and tissue penetration in mice with trinitrobenzenesulfonic acid (TNBS)-induced IBD compared to model 2 µm PS particles coated with polyvinylpyrollidone (PVP), the stabilizer used in the clinical micronized budesonide formulation. We then used a wet-milling process to develop a budesonide NS formulation with a muco-inert Pluronic F127 coating (particle size ∼230 nm), as well as a budesonide microsuspension (MS) stabilized with PVP (particle size ∼2 µm). Using an acute TNBS mouse model of IBD, we show that daily budesonide NS enema treatment resulted in a significant reduction in the macroscopic (decreased colon weight) and microscopic (histology score) symptoms of IBD compared to untreated controls or mice treated daily with the budesonide MS enema. Further, we show that the budesonide NS enema treated mice had a significantly reduced number of inflammatory macrophages and IL-ß producing CD11b + cells in colon tissue compared to untreated controls or mice treated with the budesonide MS enema. We conclude that the nano-size and muco-inert coating allowed for enhanced local delivery of budesonide, and thus, a more significant impact on local colorectal tissue inflammation.

Hypo-osmolar Formulation of Tenofovir (TFV) Enema Promotes Uptake and Metabolism of TFV in Tissues, Leading to Prevention of SHIV/SIV Infection.

Oral preexposure prophylaxis (PrEP) has been approved for prophylaxis of HIV-1 transmission but is associated with high costs and issues of adherence. Protection from anal transmission of HIV using topical microbicides and methods congruent with sexual behavior offers the promise of improved adherence. We compared the pharmacokinetics (PK) and ex vivo efficacy of iso-osmolar (IOsm) and hypo-osmolar (HOsm) rectal enema formulations of tenofovir (TFV) in rhesus macaques. Single-dose PK of IOsm or HOsm high-dose (5.28 mg/ml) and low-dose (1.76 mg/ml) formulations of TFV enemas were evaluated for systemic uptake in blood, colorectal biopsy specimens, and rectal CD4+ T cells. Markedly higher TFV concentrations were observed in plasma and tissues after administration of the HOsm high-dose formulation than with all other formulations tested. TFV and TFV diphosphate (TFV-DP) concentrations in tissue correlated for the HOsm high-dose formulation, demonstrating rapid uptake and transformation of TFV to TFV-DP in tissues. TFV-DP amounts in tissues collected at 1 and 24 h were 7 times and 5 times higher, respectively (P < 0.01), than the ones collected in tissues with the IOsm formulation. The HOsm high-dose formulation prevented infection in ex vivo challenges of rectal tissues collected at 1, 24, and 72 h after the intrarectal dosing, whereas the same TFV dose formulated as an IOsm enema was less effective.

Local enema treatment to inhibit FOLH1/GCPII as a novel therapy for inflammatory bowel disease.

Here we evaluate the potential for local administration of a small molecule FOLH1/GCPII inhibitor 2-phosphonomethyl pentanedioic acid (2-PMPA) as a novel treatment for inflammatory bowel disease (IBD). We found that FOLH1/GCPII enzyme activity was increased in the colorectal tissues of mice with TNBS-induced colitis, and confirmed that 2-PMPA inhibited FOLH1/GCPII enzyme activity ex vivo. In order to maximize local enema delivery of 2-PMPA, we studied the effect of vehicle tonicity on the absorption of 2-PMPA in the colon. Local administration of 2-PMPA in a hypotonic enema vehicle resulted in increased colorectal tissue absorption at 30min compared to 2-PMPA administered in an isotonic enema vehicle. Furthermore, local delivery of 2-PMPA in hypotonic enema vehicle resulted in prolonged drug concentrations for at least 24h with minimal systemic exposure. Finally, daily treatment with the hypotonic 2-PMPA enema ameliorated macroscopic and microscopic symptoms of IBD in the TNBS-induced colitis mouse model, indicating the potential of FOLH1/GCPII inhibitors for the local treatment of IBD.

Nanoparticles for oral delivery: Design, evaluation and state-of-the-art.

The oral route is a preferred method of drug administration, though achieving effective drug delivery and minimizing off-target side effects is often challenging. Formulation into nanoparticles can improve drug stability in the harsh gastrointestinal (GI) tract environment, providing opportunities for targeting specific sites in the GI tract, increasing drug solubility and bioavailability, and providing sustained release in the GI tract. However, the unique and diverse physiology throughout the GI tract, including wide variation in pH, mucus that varies in thickness and structure, numerous cell types, and various physiological functions are both a barrier to effective delivery and an opportunity for nanoparticle design. Here, nanoparticle design aspects to improve delivery to particular sites in the GI tract are discussed. We then review new methods for evaluating oral nanoparticle formulations, including a short commentary on data interpretation and translation. Finally, the state-of-the-art in preclinical targeted nanoparticle design is reviewed.

Natural polyphenols: potential in the prevention of sexually transmitted viral infections.

Sexually transmitted viral infections represent a major public health concern due to lack of effective prevention strategies. Efforts are ongoing to develop modalities that can enable simultaneous prevention of multiple sexually transmitted infections. In the present review, we discuss the potential of natural polyphenols to prevent sexually transmitted viral infections. The review gives an account of various in vitro and in vivo studies carried out on epigallocatechin gallate, theaflavins (black tea polyphenols), resveratrol, genistein and curcumin to highlight their potential to prevent sexually transmitted infections caused by HIV (human immunodeficiency virus), HSV (herpes simplex virus) and HPV (human papilloma virus).

Development and evaluation of a thermosensitive vaginal gel containing raltegravir+efavirenz loaded nanoparticles for HIV prophylaxis.

The objective of this investigation was to develop a thermosensitive vaginal gel containing raltegravir+efavirenz loaded PLGA nanoparticles (RAL+EFV-NPs) for pre-exposure prophylaxis of HIV. RAL+EFV-NPs were fabricated using a modified emulsion-solvent evaporation method and characterized for size and zeta potential. The average size and surface charge of RAL+EFV-NP were 81.8±6.4 nm and -23.18±7.18 mV respectively. The average encapsulation efficiency of raltegravir and efavirenz was 55.5% and 98.2% respectively. Thermosensitive vaginal gel containing RAL+EFV-NPs was successfully prepared using a combination of Pluronic F127 (20% w/v) and Pluronic F68 (1% w/v). Incorporation RAL+EFV-NPs in the gel did not result in nanoparticle aggregation and RAL+EFV-NPs containing gel showed thermogelation at 32.5°C. The RAL+EFV-NPs were evaluated for inhibition of HIV-1(NL4-3) using TZM-bl indicator cells. The EC(90) of RAL+EFV-NPs was lower than raltegravir+efavirenz (RAL+EFV) solution but did not reach significance. Compared to control HeLa cells without any treatment, RAL+EFV-NPs or blank gel were not cytotoxic for 14 days in vitro. The intracellular levels of efavirenz in RAL+EFV-NPs treated HeLa cells were above the EC(90) for 14 days whereas raltegravir intracellular concentrations were eliminated within 6 days. Transwell experiments of NPs-in-gel demonstrated rapid transfer of fluorescent nanoparticles from the gel and uptake in HeLa cells within 30 min. These data demonstrate the potential of antiretroviral NP-embedded vagina gels for long-term vaginal pre-exposure prophylaxis of heterosexual HIV-1 transmission.

Characterization of soy polysaccharide and its in vitro and in vivo evaluation for application in colon drug delivery.

The objective of the present investigation was to establish potential of commercially available soy polysaccharide (Emcosoy®) for colon drug delivery. The soy polysaccharide-ethyl cellulose films were fabricated and characterized. The effect of the pectinase enzyme on the tensile strength and surface morphology of the film was evaluated. The permeation of chlorpheniramine maleate (CPM), a model hydrophilic drug from pectinase enzyme treated and untreated films was measured in pH 7.4 buffer. The soy polysaccharide-ethyl cellulose films were also incubated with Lactobacillus sp. culture for a specific duration, and effect on the CPM permeation was evaluated. The CPM capsules were coated with the soy polysaccharide-ethyl cellulose mixture, and Eudragit S100 was applied as a secondary coat. The coated CPM capsules were radiolabelled, and their in vivo transit was evaluated in human volunteers on oral administration. The pectinase enzyme had a significant influence on the tensile strength and surface morphology of the soy polysaccharide-ethyl cellulose films. The permeability of pectinase enzyme-treated and Lactobacillus sp.-treated films was significantly higher than that of untreated films. The CPM capsules were coated with the soy polysaccharide-ethyl cellulose mixture and Eudragit S100 and were successfully radiolabelled by a simple method. Gamma scintigraphic studies in human volunteers showed that the radiolabelled capsules maintained integrity for at least 9 h after oral administration. Thus, the soy polysaccharide has a potential in colon drug delivery.

Lecithin-based novel cationic nanocarriers (LeciPlex) I: fabrication, characterization and evaluation.

AIMS: In the present investigation, the feasibility of fabricating novel self-assembled cationic nanocarriers (LeciPlex) containing cetyltrimethylammonium bromide (CTAB) or didodecyldimethylammonium bromide (DDAB) and soybean lecithin using pharmaceutically acceptable biocompatible solvents such as 2-Pyrrolidone (Soluphor P) and diethyleneglycol monoethyl ether (Transcutol) was established. MATERIALS & METHODS: The interaction between DDAB/CTAB and soybean lecithin in the nanocarriers was confirmed by differential scanning calorimetry and in vitro antimicrobial studies. The positive charge on the nanocarriers was confirmed by zeta potential analysis. RESULTS: Transmission electron microscopy analysis could not reveal sufficient information regarding the internal structure of the nanocarriers, whereas cryotransmission electron microscopy studies indicated that these novel nanocarriers have unilamellar structure. Small-angle neutron scattering studies confirmed interaction of cationic surfactant (DDAB) and lecithin in the nanocarriers and confirmed the presence of unilamellar nanostructures. CONCLUSION: Various hydrophobic drugs could be encapsulated in the CTAB/DDAB-based lecithin nanocarriers (CTAB-LeciPlex or DDAB-LeciPlex) irrespective of their difference in log p-values. In vitro antimicrobial studies on triclosan-loaded LeciPlex confirmed entrapment of triclosan in the nanocarriers. The ability of CTAB-LeciPlex and DDAB-LeciPlex to condense plasmid DNA was established using agarose gel electrophoresis. DDAB-LeciPlex could successfully transfect pDNA in HEK-293 cells indicating potential in gene delivery.

Lecithin-based novel cationic nanocarriers (Leciplex) II: improving therapeutic efficacy of quercetin on oral administration.

The objective of the present investigation was to evaluate ability of the novel self-assembled phospholipid- based cationic nanocarriers (LeciPlex) in improving the therapeutic efficacy of a poorly water-soluble natural polyphenolic agent, quercetin (QR), on oral administration. Quercetin loaded LeciPlex (QR-LeciPlex) were successfully fabricated using a biocompatible solvent Transcutol HP. The QR-LeciPlex were characterized for particle size, encapsulation efficiency, zeta potential, and particle morphology by cryo-TEM. UV and fluorescence spectral characterization was carried out to find out the association of QR with LeciPlex. Small angle neutron scattering studies (SANS) were carried out to understand the internal structure of Leciplex and to evaluate the influence of the incorporation of QR in the LeciPlex. Anti-inflammatory and antitumorigenic activity of QR-LeciPlex was determined in comparison to QR suspension to evaluate the potential of LeciPlex in improving oral delivery of QR. QR-LeciPlex exhibited a particle size of ∼400 nm and had excellent colloidal stability. The QR-LeciPlex had a zeta potential greater than +30 mV and exhibited very high encapsulation efficiency of QR (>90%). UV and fluorescence spectral characterization indicated the interaction/association of QR with LeciPlex components. Cryo-TEM studies showed that LeciPlex and QR-LeciPlex have a unilamellar structure. SANS confirmed the unilamellar structure of LeciPlex and indicated that the incorporation of QR does not have any effect on the internal structure of the LeciPlex. QR-LeciPlex exhibited significantly higher anti-inflammatory and antitumorigenic activity (p < 0.01) as compared to that of QR suspension on oral administration.

Design and evaluation of microemulsions for improved parenteral delivery of propofol.

The objective of this investigation was to evaluate the potential of the microemulsions to improve the parenteral delivery of propofol. Pseudo-ternary phase diagrams were plotted to identify microemulsification region of propofol. The propofol microemulsions were evaluated for globule size, physical and chemical stability, osmolarity, in vitro hemolysis, pain caused by injection using rat paw-lick test and in vivo anesthetic activity. The microemulsions exhibited globule size less than 25 nm and demonstrated good physical and chemical stability. Propofol microemulsions were slightly hypertonic and resulted in less than 1% hemolysis after 2 h of storage with human blood at 37 degrees C. Rat paw-lick test indicated that propofol microemulsions were significantly less painful as compared to the marketed propofol formulation. The anesthetic activity of the microemulsions was similar to the marketed propofol formulation indicating that they do not compromise the pharmacological action of propofol. The stability studies indicated that the microemulsions were stable for 3 months when stored at 5 +/- 3 degrees C. Thus, microemulsions appeared to be an interesting alternative to the current propofol formulations.