Myung Joo Kang

Surface modification of lipid-based nanocarriers for cancer cell-specific drug targeting

Targeted drug delivery systems using nanocarriers for anticancer drugs have been investigated for over several decades. Among the many nanocarrier systems, lipid-based nanocarriers such as liposomes, solid lipid nanoparticles, and nanostructured lipid carriers have afforded attention as a carrier system to improve the efficacy of anticancer drugs. Recent efforts have focused on cancer cell-specific drug delivery through the functionalization of the surface of lipid-based nanocarriers with various ligands such as targeting moieties, cell-penetrating peptides, and cell-penetrating homing peptides to overcome non-selectivity, minimize side effects, and enhance antitumor efficacy. However, the use of ligand modification has been limited because the nanocarriers were easily recognized by the mononuclear phagocyte system and thus rapidly removed from the blood circulation. To achieve prolonged systemic circulation, nanocarriers were further modified with protective polymers such as polyethylene glycol (PEG). Unexpectedly, this presented a PEG dilemma, as the interaction of ligands with the target was hindered and induced poor cellular uptake. Recently, stimuli-sensitive cleavage of the PEG coat, following recognition of the cancer cell microclimate, such as low pH, redox-potential, and over-expressed enzymes, was established to solve this problem. This review presents a comprehensive overview on the current state of surface-modified lipid-based nanocarriers for the improved delivery of anticancer drugs.

RIPL peptide (IPLVVPLRRRRRRRRC)-conjugated liposomes for enhanced intracellular drug delivery to hepsin-expressing cancer cells

BACKGROUND To facilitate selective drug delivery to hepsin (Hpn)-expressing cancer cells, the RIPL peptide (IPLVVPLRRRRRRRRC; 16mer; 2.1 kDa) was synthesized as a novel cell penetrating/homing peptide (CPHP) and conjugated to a liposomal carrier. METHODS RIPL peptide-conjugated liposomes (RIPL-Lipo) were prepared by conjugating RIPL peptides to maleimide-derivatized liposomal vesicles via the thiol-maleimide reaction. Vesicle size and zeta potential were examined using a Zetasizer. Intracellular uptake specificity of the RIPL peptide, or RIPL-Lipo, was assessed by measuring mean fluorescence intensity (MFI) after treatment with a fluorescent marker in various cell lines: SK-OV-3, MCF-7, and LNCaP for Hpn(+); DU145, PC3, and HaCaT for Hpn(-). FITC-dextran was used as a model compound. Selective translocational behavior of RIPL-Lipo to LNCaP cells was visualized by fluorescence microscopy and confocal laser scanning microscopy. Cytotoxicities of the RIPL peptide and RIPL-Lipo were evaluated by WST-1 assay. RESULTS RIPL peptides exhibited significant Hpn-selectivity. RIPL-Lipo systems were of positively charged nanodispersion (165 nm in average; 6-24 mV depending on RIPL conjugation ratio). RIPL-Lipo with the conjugation of 2300 peptide molecules revealed the greatest MFI in all cell lines tested. Cellular uptake of RIPL-Lipo increased by 20- to 70-fold in Hpn(+) cells, and 5- to 7-fold in Hpn(-) cells, compared to the uptake of FITC-dextran. Cytosolic internalization of RIPL-Lipo was time-dependent: bound instantly; internalized within 30 min; distributed throughout the cytoplasm after 1 h. Cytotoxicities of RIPL peptide (up to 50 μM) and RIPL-Lipo (up to 10%) were minor (cell viability >90%) in LNCaP and HaCaT cells. CONCLUSION By employing a novel CPHP, the RIPL-Lipo system was successfully developed for Hpn-specific drug delivery.

Improved oral absorption of dutasteride via Soluplus®-based supersaturable self-emulsifying drug delivery system (S-SEDDS).

A novel supersaturable self-emulsifying drug delivery system (S-SEDDS) was formulated to improve the oral absorption of dutasteride (DTS), a 5α-reductase inhibitor that is poorly water-soluble. A supersaturable system was prepared by employing Soluplus(®) (polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer) as a precipitation inhibitor with a conventional SEDDS vehicle consisted of Capryol™ 90, Cremophor(®) EL and Transcutol(®) HP (DTS:SEDDS vehicle:Soluplus(®)=1.0:67.6:10.0 w/v/w). In an in vitro dissolution test in a non-sink condition, the drug dissolution rate from SEDDS was rapidly increased to 72% for an initial period of 5min, but underwent rapid drug precipitation within 2h, decreasing the amount of drug dissolved to one-seventh of its original amount. On the other hand, S-SEDDS resulted in a slower crystallization of DTS by virtue of a precipitation inhibitor, maintaining a 3 times greater dissolution rate after 2h compared to SEDDS. In an in vivo pharmacokinetic study in rats, the S-SEDDS formulation exhibited 3.9-fold greater area-under-curve value than that of the drug suspension and 1.3-fold greater than that of SEDDS. The maximum plasma concentration of S-SEDDS was 5.6- and 2.0-fold higher compared to drug suspension and SEDDS, respectively. The results of this study suggest that the novel supersaturable system may be a promising tool for improving the physicochemical property and oral absorption of the 5α-reductase inhibitor.

Improved oral absorption and chemical stability of everolimus via preparation of solid dispersion using solvent wetting technique

The aim of this study was to improve the physicochemical properties and oral absorption of poorly water-soluble everolimus via preparation of a solid dispersion (SD) system using a solvent wetting (SW) technique. The physicochemical properties, drug release profile, and bioavailability of SD prepared by SW process were also compared to SD prepared by the conventional co-precipitation method. Solid state characterizations using scanning electron microscopy, particle size analysis and X-ray powder diffraction indicated that drug homogeneously dispersed and existed in an amorphous state within the intact polymeric carrier. Whereas, a film-like mass was obtained by a co-precipitation method and further pulverization step was needed for tabletization. The drug release from the SD tablet prepared by SW process at a ratio of drug to hydroxypropyl methylcellulose of 1:15 was markedly higher than the drug alone and equivalent to the marketed product (Afinitor(®), Novartis Pharmaceuticals), a SD tablet prepared by co-precipitation method, archiving over 75% the drug release after 30 min. At the accelerated (40°C/75% R.H.) and stress (80°C) stability tests, the novel formula was more stable than drug powder and provided comparable drug stability with the commercially available product, which contains a potentially risky antioxidant, butylated hydroxyl toluene. The pharmacokinetic parameters after single oral administration in beagles showed no significant difference (P>0.01) between the novel SD-based tablet and the marketed product. The results of this study, therefore, suggest that the novel SD system prepared by the solvent wetting process may be a promising approach for improving the physicochemical stability and oral absorption of the sirolimus derivatives.

Improved oral absorption of tacrolimus by a solid dispersion with hypromellose and sodium lauryl sulfate

A novel surfactant-incorporated hydroxypropyl methylcellulose (HPMC) solid dispersion (SD) system was constructed in order to facilitate the release rate and oral absorption of tacrolimus (FK506), a poorly water-soluble immunosuppressant. Several emulsifiers including sodium lauryl sulfate (SLS), as drug release promotors, were employed with HPMC to fabricate SD using the solvent wetting method. The solid state characteristics using differential scanning calorimetry and X-ray powder diffraction, revealed that FK506 was molecularly distributed within all dispersions in amorphous form. The dissolution rates of FK506 in SLS-incorporated SDs were much higher than those in SDs prepared with HPMC alone, and even with stearoyl polyoxyl-32 glycerides or tocopheryl polyethylene glycol 1000 succinate. In particular, the greatest dissolution enhancement was obtained from the SD consisting of the drug, HPMC, and SLS in a weight ratio of 1:1:3, providing a 50-fold higher drug concentration within 15 min, compared with HPMC SD. In vivo absorption study in rats demonstrates that the optimized formula remarkably increased the oral absorption of FK506, providing about 4.0-fold greater bioavailability (p<0.05) compared with the marketed product (Prograf®, Astellas Pharma). These data suggest that a novel SLS/HPMC SD may be an advantageous dosage form of FK506, boosting the dissolution and absorption in gastrointestinal tract.

Comparison of Piroxicam Pharmacokinetics and Anti-Inflammatory Effect in Rats after Intra-Articular and Intramuscular Administration

This study evaluated the pharmacokinetic profile and therapeutic efficacy of piroxicam (PX), a long acting non-steroidal anti-inflammatory drug for the treatment of arthritis, following intra-articular (IA) injection in comparison to the pharmacokinetic profile and therapeutic efficacy of PX after intramuscular (IM) injection. In the pharmacokinetic study in rats, systemic exposure and pharmacokinetic parameters of PX after a single IA dose were compared with systemic exposure and pharmacokinetic parameters of PX after administration of the same dose IM (0.6 mg/kg). The anti-inflammatory and analgesic effects of IA PX were evaluated simultaneously in a monoiodoacetate-induced osteoarthritis rat model. The plasma PX concentration rapidly rose following IA injection, and it was comparable to the plasma PX concentration following IM injection, suggesting the rapid efflux of the drug molecule from the joint cavity. However, in the efficacy study, the IA PX administration significantly reduced the knee swelling by reducing the level of prostaglandin E2 in the joint, compared to that following administration of IA vehicle and after administration of the IM PX dose. In addition, we found that the anti-inflammatory and anti-nociceptive efficacies of IA PX were synergistically increased upon co-treatment with hyaluronic acid (HA), a potent agent for the treatment of osteoarthritis, at the weight ratio of 1:1 or 1:2, and these effects were more pronounced than those following administration of HA or PX alone. In conclusion, this study demonstrated the efficacy of the IA use of PX alone and/or in combination with HA in osteoarthritis.

Design of pH-responsive alginate raft formulation of risedronate for reduced esophageal irritation

Risedronate sodium (RA), a pyridinyl bisphosphonate, is widely used in the treatment of osteoporosis. However, the free acid form of the bisphosphonate below pH 3.5 has the potential to produce severe impatience of the upper gastrointestinal tract, particularly esophagitis. A pH-responsive raft-forming tablet (PRR-T) was designed to prevent the esophageal irritation, mainly consisting of low-molecular-weight alginate (LFR 5/60, 300 mg) as raft-forming polymer, sodium bicarbonate (1000 mg) as gas-generating agent and citrate and sodium citrate (600 and 200 mg, respectively) as buffer system. A PRR-T was rapidly liquefied in water within 80 s with a low viscosity 8.0 mPa s, offering ease of swallowing in patients. A formulation profoundly neutralized simulated gastric fluid over pH 5.5, leading to an ionization of the bisphosphonate, without raft formation. On the other hand, the raft was rapidly formed on the top layer preventing the reflux of RA, if the contact with acidic medium is much higher than 0.5 N of hydrochloric acid. Nevertheless, the release rate of the drug was equivalent, providing over 95% release within 5 min. Our study demonstrated the potential usefulness of alginate-based PRR-T for an oral therapy with bisphosphonates for reduced esophageal adverse experiences.

Formulation and in vivo evaluation of probiotics-encapsulated pellets with hydroxypropyl methylcellulose acetate succinate (HPMCAS).

The aim of this study was to formulate probiotics-encapsulated pellets with hydroxypropyl methylcellulose acetate succinate (HPMCAS) using a dry powder coating technique to improve the storage stability, acid resistance, and intestinal adherence of viable bacteria (Lactobacillus acidophilus and Bifidobacteria animalis ssp. Lactis). Dry coated pellet (DCP) loaded with probiotics was optimized with respect to the quantity of the HPMCAS, an enteric coating polymer (108 mg), and the kinds and amounts of plasticizer (triethyl citrate, 15.7 mg; acetylated monoglyceride, 6.8 mg), by evaluating the survival rate of the bacteria during preparation process and in an acidic medium. Dry coating process allows the whole survivals of living bacteria during preparation process. The DCP formulation exhibited markedly higher acid tolerability and storage stability compared to uncoated viable bacteria. In an in vivo mucosal adherence study in rats, a profound colonization of viable bacteria in the small and large intestine was observed in rats receiving DCP system (p<0.05) compared to rats receiving uncoated probiotics. Moreover, we found that the repeated DCP administration noticeably inhibited intestinal penetration of endotoxin, a potent inflammatory stimulant, from intestinal mucus. The novel DCP system may be an alternative approach for improving bacterial viability in the preparation process and in an acidic medium, and to promote mucosal colonization of probiotic bacteria in the human gut.

Reduced Food-Effect on Intestinal Absorption of Dronedarone by Self-microemulsifying Drug Delivery System (SMEDDS)

The oral absorption of dronedarone (DRN), a benzofuran derivative with anti-arrhythmic activity, is significantly affected by food intake. The absolute bioavailability of the marketed product (Multaq, Sanofi, U.S.) was about 4% without food, but increased to 15% when administered with a high fat meal. Therefore, to reduce the food-effect on the intestinal absorption of DRN, a novel self-microemulsifying drug delivery system (SMEDDS) was formulated and the comparative in vivo absorption studies with the marketed product were carried out using male beagle dogs either in the fasted or fed state. The SMEDDS consisted of the drug, Labrafil M 1944CS, and Kolliphor EL in a weight ratio of 1 : 1 : 2, rapidly formed a fine oil-in-water emulsion with a droplet size less than 50 nm. An in vivo absorption study revealed that the area-under-curve (AUC0-24 h) and maximal plasma concentration (Cmax) were 10.4-fold (p<0.05) and 8.6-fold (p<0.05) higher, respectively, after the marketed product was orally administered to beagles in the fed state when compared to those in the fasted state. This food-effect were remarkably alleviated by SMEDDS formulation, with AUC0-24 h and Cmax 2.9-fold (p<0.05) and 2.6-fold (p<0.05) higher in the fed state when compared to the fasted state, by facilitating intestinal absorption of DRN in the fasted state. The results of this study suggest that SMEDDS may decrease the differences in oral absorption of DRN between the prandial states, improving therapeutic efficacy as well as patient compliance.

Docetaxel-loaded RIPL peptide (IPLVVPLRRRRRRRRC)-conjugated liposomes: Drug release, cytotoxicity, and antitumor efficacy

We previously synthesized the RIPL peptide (IPLVVPLRRRRRRRRC) to facilitate selective delivery into hepsin-expressing cancer cells and showed that RIPL peptide-conjugated liposomes (RIPL-L) enhanced the intracellular delivery of fluorescent probes in vitro. In this study, docetaxel-loaded RIPL-L (DTX-RIPL-L) were prepared and evaluated for in vitro drug release, cytotoxicity, and in vivo antitumor efficacy. DTX was successfully encapsulated by pre-loading, with an average encapsulation efficiency and drug loading capacity of 32.4% and 21.39±2.05 (μg/mg), respectively. A DTX release study using dialysis showed a biphasic release pattern, i.e., rapid release for 6h, followed by sustained release up to 72h. The first-order equation provided the best fit for drug release (r2=0.9349). In vitro cytotoxicity was dose-dependent, resulting in IC50 values of 36.10 (SK-OV-3) and 48.62ng/mL (MCF-7) for hepsin-positive, and 61.12 (DU145) and 53.04ng/mL (PC-3) for hepsin-negative cell lines. Live/dead cell imaging was carried out to visualize the proportion of viable and nonviable SK-OV-3 cells. Compared to DTX solution, DTX-RIPL-L significantly inhibited tumor growth and prolonged survival time in BALB/c nude mice with SK-OV-3 cell tumors. We suggest that DTX-RIPL-L is a good candidate for efficient drug targeting to hepsin-expressing cancer cells.