The complexity of intestinal absorption and exsorption of digoxin in rats

The potential multiple carrier-mediated mechanisms involved in the transport of digoxin in rat intestine were investigated by the rapid filtration method in rat intestinal brush-border vesicles (BBMV) and in vitro Ussing chambers. The uptake of digoxin showed a typical overshoot phenomenon in the presence of an inward proton gradient and an outward bicarbonate gradient, or an outward glutathione gradient in BBMV. Good fitting to an equation consisting of both saturable and linear terms was obtained using non-linear regression analysis. GF120918, a specific P-gp inhibitor, significantly increased the absorptive permeability of digoxin in rat ileum (7.02 x 10(-7) cm/s versus 2.11 x 10(-6) cm/s with GF120918) but the addition of DIDS (0.5 mM), an anionic transporter inhibitor, or bromosulfophthalein (0.1 mM), an Oatp inhibitor, in the presence of GF120918 decreased the absorptive permeability compared with GF120918 alone (2.11 x 10(-6) cm/s versus 1.46 x 10(-6) cm/s, p<0.01 and 2.11 x 10(-6) cm/s versus 1.60 x 10(-6) cm/s, p<0.05, respectively). The above results suggest the involvement of carrier-mediated uptake mechanism, possibly Oatp, in digoxin absorption. Interestingly, GF120918 (1 microM) did not abolish the polarized transport of digoxin in rat jejunum and ileum, and DIDS (0.5 mM), not a P-gp inhibitor, and MK571 (50 microM), an MRP-selective inhibitor, can also significantly decrease the exsorptive permeability of digoxin. This result indicates the involvement of non-P-gp efflux transporter in digoxin secretion and this transporter is DIDS and MK571-sensitive. Contrary to conventional concept, our studies show that intestinal absorption of digoxin may involve both active uptake and efflux transporters. Our study may have clinical implications in drug-drug or drug-food interactions involving transporters.     

Efficacy of novel P-glycoprotein inhibitors to increase the oral uptake of paclitaxel in mice

P-glycoprotein inhibitors can increase the oral bioavailability of paclitaxel. We have now explored the mechanisms that determine the efficacy of several novel P-glycoprotein inhibitors to increase the absorption of paclitaxel from the gut lumen of mice in both in vivo and in vitro experiments. The inhibitors studied were cyclosporin A, PSC 833, GF120918, LY335979 and R101933. Mass balance studies showed that GF120918 was the most effective inhibitor, resulting in almost complete uptake of paclitaxel. PSC 833 was slightly less effective, whereas cyclosporin A and LY335979 were moderately effective. R101933 had only marginal effects. These findings were in line with in vitro transport experiments using LLC-mdr1a cells. By studying the intra-intestinal kinetics of the agents we found that cyclosporin A, PSC 833 and GF120918 rapidly passed the stomach and traveled concurrently with paclitaxel through the intestines, whereas LY335979 and R101933 delayed stomach emptying. Moreover, these latter compounds appear to be more readily absorbed when released into the intestines thus reducing local intestinal concentrations. Due to their combined effects on absorption and metabolic elimination of paclitaxel, cyclosporin A and PSC 833 resulted in the highest paclitaxel levels in plasma. In conclusion, our models provide insight into the factors that determine the suitability of P-glycoprotein inhibitors to enable oral paclitaxel therapy and will be useful in selecting candidate inhibitors for clinical testing.     

Preparation and in vivo evaluation of SMEDDS (self-microemulsifying drug delivery system) containing fenofibrate

The present work was aimed at formulating a SMEDDS (self-microemulsifying drug delivery system) of fenofibrate and evaluating its in vitro and in vivo potential. The solubility of fenofibrate was determined in various vehicles. Pseudoternary phase diagrams were used to evaluate the microemulsification existence area, and the release rate of fenofibrate was investigated using an in vitro dissolution test. SMEDDS formulations were tested for microemulsifying properties, and the resultant microemulsions were evaluated for clarity, precipitation, and particle size distribution. Formulation development and screening was done based on results obtained from phase diagrams and characteristics of resultant microemulsions. The optimized formulation for in vitro dissolution and pharmacodynamic studies was composed of Labrafac CM10 (31.5%), Tween 80 (47.3%), and polyethylene glycol 400 (12.7%). The SMEDDS formulation showed complete release in 15 minutes as compared with the plain drug, which showed a limited dissolution rate. Comparative pharmacodynamic evaluation was investigated in terms of lipid-lowering efficacy, using a Triton-induced hypercholesterolemia model in rats. The SMEDDS formulation significantly reduced serum lipid levels in phases I and II of the Triton test, as compared with plain fenofibrate. The optimized formulation was then subjected to stability studies as per International Conference on Harmonization (ICH) guidelines and was found to be stable over 12 months. Thus, the study confirmed that the SMEDDS formulation can be used as a possible alternative to traditional oral formulations of fenofibrate to improve its bioavailability.     

Influence of Passive Permeability on Apparent P-glycoprotein Kinetics

Purpose. The objectives of this work were to evaluate the importance of moderate passive permeability on apparent P-glycoprotein (P-gp) kinetics, and demonstrate that inspection of basolateral to apical and apical to basolateral (BL-AP/AP-BL) permeability ratios may result in a compound being overlooked as a P-gp substrate and inhibitor of another drug's transport via P-gp inhibition. Methods. The permeability ratios of nicardipine, vinblastine, cimetidine, and ranitidine were determined across Caco-2 monolayers that express P-gp, in the presence and absence of the specific P-gp inhibitor, GF120918. In addition, the permeability ratio of vinblastine was studied after pretreatment of Caco-2 monolayers with nicardipine, ranitidine, or cimetidine. Similar studies were repeated with hMDR1-MDCK monolayers. Results. The permeability ratios for cimetidine and vinblastine were >2. The permeability ratios for nicardipine and ranitidine were close to unity, and were not affected by the addition of GF120918. Based solely on ratios, only compounds with moderate transcellular permeability (vinblastine and cimetidine) would be identified as P-gp substrates. Although the permeability ratios appeared to be unity for nicardipine and ranitidine, both compounds affected the permeability of vinblastine, and were identified as substrates and inhibitors of P-gp. Studies performed in hMDR1-MDCK cells confirmed these experimental results. Data were explained in the context of a kinetic model, where passive permeability and P-gp efflux contribute to overall drug transport. Conclusions. Moderate passive permeability was necessary for P-gp to reduce the AP-BL drug permeability. Inspection of the permeability ratio after directional transport studies did not effectively identify P-gp substrates that affected the P-gp kinetics of vinblastine. Because of the role of passive permeability, drug interaction studies with known P-gp substrates, rather than directional permeability studies, are needed to elucidate a more complete understanding of P-gp kinetics.     

Enhancement of solubility and therapeutic potential of poorly soluble lovastatin by SMEDDS formulation adsorbed on directly compressed spray dried magnesium aluminometasilicate liquid loadable tablets: A study in diet induced hyperlipidemic rabbits

The aim of present study was to formulate and evaluate a self-microemulsifying drug delivery systems (SMEDDS) containing lovastatin and to further explore the ability of porous Neusilin^® US2 tablet as a solid carrier for SMEDDS. SMEDDS formulations of varying proportions of peceol, cremophor RH 40 and transcutol-P were selected and subjected to in-vitro evaluation, including dispersibility studies, droplet size, zeta potential measurement and release studies. The results indicated that the drug release profile of lovastatin from SMEDDS formulations was statistically significantly higher (p-value < 0.05) than the plain lovastatin powder. Thermodynamic stability studies also confirmed the stability of the prepared SMEDDS formulations. The optimized formulation, which consists of 12% of peceol, 44% of cremophor RH 40, and 44% of transcutol-P was loaded into directly compressed liquid loadable tablet of Neusilin^® US2 by simple adsorption method. In order to determine the ability of Neusilin^® US2 as a suitable carrier pharmacodynamics study were also carried out in healthy diet induced hyperlipidemic rabbits. Animals were administered with both liquid SMEDDS and solid SMEDDS as well. From the results obtained, Neusilin^® was found to be a suitable carrier for SMEDDS and was equally effective in reducing the elevated lipid profile. In conclusion, liquid loadable tablet (LLT) is predicted to be a promising technique to deliver a liquid formulation in solid state.     

Drug efflux transport properties of 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester (BCECF-AM) and its fluorescent free acid, BCECF

2',7'-Bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) is a fluorescent probe used to examine multidrug resistance-associated protein (MRP) transporter activity in cells. BCECF is introduced into the cell as the nonfluorescent membrane permeable acetoxymethyl ester, BCECF-AM, where it is hydrolyzed to the membrane impermeable BCECF. The lipophilic nature of BCECF-AM suggests it may be a substrate for other drug efflux transporters such as P-glycoprotein (P-gp) and the breast cancer resistance protein (BCRP). To assess the drug efflux transporter interactions of BCECF-AM and BCECF, accumulation studies were examined in various drug efflux-expressing cells. Inhibition of P-gp, BCRP, and/or MRP produced distinct changes in the time-dependent accumulation of BCECF in the cells. Treatment with GF120918 produced an immediate and sustained effect throughout the entire time course examined. Fumitremorgin C only affected BCECF accumulation at the early time points, whereas the impact of indomethacin on BCECF accumulation was observed only at the latter time points. Permeability studies in bovine brain microvessel endothelial cells indicated an increased basolateral-to-apical transport of BCECF, which could be reduced in the presence of either indomethacin or GF120918. These results indicate that the intracellular accumulation and transcellular permeability of BCECF are sensitive to a variety of drug efflux interactions. These results likely reflect an interaction of the ester form with P-gp and BCRP during the initial accumulation process, and an interaction of the free acid form with MRP after hydrolysis in the cell.     

Effect of self-microemulsifying drug delivery systems containing Labrasol on tight junctions in Caco-2 cells

The aim of this study was to investigate the effect of two novel self-microemulsifying drug delivery systems (SMEDDS) containing Labrasol with different dilutions on tight junctions. Changes in barrier properties of Caco-2 cell monolayers, including transepithelial electrical resistance (TEER) and permeability to the paracellular marker, i.e., mannitol, were assessed in response to dilutions and surfactants contents within formulations. The cytotoxicity of SMEDDS and the effect of surfactants on Caco-2 cells were evaluated by the MTT. Changes in subcellular localization of the tight junction proteins, ZO-1 and F-actin, were examined by confocal laser scanning microscopy. Results demonstrated that negatively charged SMEDDS with different dilutions had no effect on the TEER, but significantly increased the permeability of mannitol. In contrast, the positively charged formulation showed a dilution-dependent reduction in TEER. A corresponding increase in mannitol permeability of up to 29.4-fold to 64.7-fold greater than the control was also observed across the monolayer. Labrasol with the concentration of 0.1 and 1% was shown to increase the permeability of mannitol by 4.6-fold and 33.8-fold, respectively. The mechanism of opening of tight junctions was found to involve F-actin-related changes and redistribution of ZO-1.     

Self-microemulsifying drug delivery system (SMEDDS) improves anticancer effect of oral 9-nitrocamptothecin on human cancer xenografts in nude mice.

9-Nitrocamptothecin (9-NC) is an orally administered topoisomerase-I inhibitor for the treatment of pancreatic carcinoma, but its oral absorption and bioavailability are poor. The main objective of this study was to develop optimal 9-nitrocamptothecin (9-NC) microemulsion prepared by self-microemulsifying drug delivery system (SMEDDS). Two SMEDDS formulations of 9-NC prepared from a mixture of ethyl oleate, Tween-80 (T-form) or Cremophor EL (C-form), and PEG-400/ethanol were formed as microemulsions under dilution with aqueous phase. The resulting microemulsions were evaluated in vitro and in vivo, including the kinetics and antitumor effects in SKOV-3 human ovarian cancer xenograft in nude mice. Following 1:10 aqueous dilution of optimal 9-NC SMEDDS, the droplet sizes of resulting microemulsions were (30.8+/-4.6)nm and (39.8+/-8.2)nm for SMEDDS T-form and C-form, respectively, and the zeta potential values were -(4.3+/-0.5)mV and -(5.7+/-0.5)mV, respectively. In SKOV-3 cells, the growth inhibition (IC(50)) of various 9-NC formulations was greatest with SMEDDS T-form (3.5+/-0.7nM) followed by SMEDDS C-form (4.6+/-0.4nM), 9-NC solution (6.6+/-1.4nM) and 9-NC suspension (26.0+/-2.9nM) (P<0.01). It was indicated that the area under the plasma concentration-time curve (AUC(0-->8h)) values of various formulations of 9-NC after oral administration ranked as the following sequence: SMEDDS T-form (360.12+/-19.44ngh/ml) approximately SMEDDS C-form (351.71+/-33.66ngh/ml)>9-NC solution (241.21+/-24.67ngh/ml)>9-NC suspension (161.24+/-24.31ngh/ml). The 9-NC SMEDDS formulations also produced significantly more tumor shrinkage (P<0.01) when compared to 9-NC suspension in nude mice bearing human ovarian cancer xenografts. The results suggest that SMEDDS is a promising drug delivery system to increase the oral bioavailability and antitumor effects of 9-NC and may be applied to other lipophilic drugs. 9-NC SMEDDS represents a novel 9-NC therapy for cancer patients.     

In‐vitro respiratory drug absorption models possess nominal functional P‐glycoprotein activity

Objectives The P‐glycoprotein (P‐gp) efflux pump is known to be present within several major physiological barriers including the brain, kidney, intestine and placenta. However, the function of P‐gp in the airways of the lung is unclear. The purpose of this study was to use the highly specific P‐gp inhibitor GF120918A to investigate the activity of the P‐gp transporter in the airways to determine whether P‐gp could influence inhaled drug disposition. Methods P‐gp activity was measured as a change in digoxin transport in the presence of GF120918A in normal human bronchial epithelial (NHBE) cells, Calu‐3 cell layers and the ex‐vivo rat lung. Key findings The efflux ratios (ERs) in NHBE and Calu‐3 cells were between 0.5 and 2, in contrast to 10.7 in the Caco‐2 cell control. These low levels of GF120918A‐sensitive polarised digoxin transport were measured in the absorptive direction in NHBE cells (ER = 0.5) and in the secretory direction in Calu‐3 cells (ER = 2), but only after 21 days in culture for both cell systems and only in Calu‐3 cells at passage >50. The airspace to perfusate transfer kinetics of digoxin in the ex‐vivo rat lung were unchanged in the presence of GF120918A. Conclusions These results demonstrated that although low levels of highly culture‐dependent P‐gp activity could be measured in cell‐lines, these should not be interpreted to mean that P‐gp is a major determinant of drug disposition in the airways of the lung.     

Development and evaluation of self-microemulsifying liquid and granule formulations of Brucea javanica oil

The aim of this study was to develop and characterize a self-microemulsifying drug delivery system (SMEDDS) of Brucea javanica oil (BJO) and transform the liquid formulation into solid granules. Solubility studies of BJO and pseudo-ternary phase diagrams were used to identify the most efficient self-emulsification region. A methyl thiazolyl tetrazolium (MTT) assay was performed to identify cell apoptosis. Antitumor activity studies were also employed to evaluate the BJO SMEDDS. The optimized BJO SMEDDS in liquid and granule formulations rapidly formed fine oil-in-water microemulsions with particle sizes <50 nm. Additionally, the MTT assay demonstrated that BJO SMEDDS had a significant effect on cancer cells, and antitumor activity studies showed remarkable inhibition of S180 tumors. The BJO SMEDDS, optimized to have good characteristics, was successfully transformed into solid granules by adsorbing onto crospovidone. The studies of the release of the BJO SMEDDS of liquid and granules in vitro suggested that the release of BJO was enhanced by the SMEDDS. These studies revealed that the new self-microemulsifying systems of liquid and granule forms might be promising strategies for the oral delivery of the poorly water-soluble drug BJO.     

Self-microemulsifying drug delivery system (SMEDDS) of vinpocetine: formulation development and in vivo assessment

A new self-microemulsifying drug delivery system (SMEDDS) has been developed to increase the solubility, dissolution rate and oral bioavailability of vinpocetine (VIP), a poor water-soluble drug. The formulations of VIP-SMEDDS were optimized by solubility assay, compatibility tests, and pseudo-ternary phase diagrams analysis. The optimal ratio in the formulation of SMEDDS was found to be Labrafac : oleic acid : Cremophor EL : Transcutol P=40 : 10 : 40 : 10 (w/w). The average particle diameter of VIP was less than 50 nm. In vitro dissolution study indicated that the dialysis method in reverse was better than the ultrafiltration method and the dialysis method in simulating the drug in vivo environment. Comparing with VIP crude drug power and commercial tablets, (-)VIP-SMEDDS caused a 3.4- and 2.9-fold increase in the percent of accumulated dissolution at 3 h. Further study on the absorption property of VIP-SMEDDS employing in situ intestine of rats demonstrated that VIP in SMEDDS could be well-absorbed in general intestinal tract without specific absorption sites. In addition, the developed SMEDDS formulations significantly improved the oral bioavailability of VIP in rats. Relative bioavailability of (-)VIP-SMEDDS and (+)VIP-SMEDDS increased by 1.85- and 1.91-fold, respectively, in relative of VIP crude powder suspension. The mechanisms of enhanced bioavailability of VIP might contribute to the improved release, enhanced lymphatic transport, and increased intestinal permeability of the drug.     

Integration of Preclinical and Clinical Data with Pharmacokinetic Modeling and Simulation to Evaluate Fexofenadine as a Probe for Hepatobiliary Transport Function

Purpose  The suitability of fexofenadine as a probe substrate to assess hepatobiliary transport function in humans was evaluated by pharmacokinetic modeling/simulation and in vitro/in situ studies using chemical modulators. Methods  Simulations based on a pharmacokinetic model developed to describe fexofenadine disposition in humans were conducted to examine the impact of altered hepatobiliary transport on fexofenadine disposition. The effect of GF120918 on fexofenadine disposition was evaluated in human sandwich-cultured hepatocytes (SCH). Additionally, the effect of GF120918, bosentan, and taurocholate on fexofenadine disposition in perfused livers from TR⁻ Wistar rats was examined. Results  Based on modeling/simulation, fexofenadine systemic exposure was most sensitive to changes in the hepatic uptake rate constant, and did not reflect changes in hepatic exposure due to altered hepatic efflux. GF120918 did not impair fexofenadine biliary excretion in human SCH. GF120918 coadministration significantly decreased Cl’_biliary to 27.5% of control in perfused rat livers. Conclusions  Simulations were in agreement with perfused liver data which predicted changes in fexofenadine systemic exposure primarily due to altered hepatic uptake. Fexofenadine is not a suitable probe to assess hepatic efflux function based on systemic concentrations. GF120918-sensitive protein(s) mediate fexofenadine biliary excretion in rat liver, whereas in human hepatocytes multiple efflux proteins are involved in fexofenadine hepatobiliary disposition.     

P-glycoprotein-mediated transport of morphine in brain capillary endothelial cells.

Cell accumulation, transendothelial permeability, and efflux studies were conducted in bovine brain capillary endothelial cells (BBCECs) to assess the role of P-glycoprotein (P-gp) in the blood-brain barrier (BBB) transport of morphine in the presence and absence of P-gp inhibitors. Cellular accumulation of morphine and rhodamine 123 was enhanced by the addition of the P-gp inhibitors N-{4-[2-(1,2,3,4-tetrahydro-6,7dimethoxy-2-isoquinolinyl)-ethyl]-phenyl}-9,10-dihydro-5-methoxy-9- carboxamide (GF120918), verapamil, and cyclosporin A. Positive (rhodamine 123) and negative (sucrose and propranolol) controls for P-gp transport also were assessed. Morphine glucuronidation was not detected, and no alterations in the accumulation of propranolol or sucrose were observed. Transendothelial permeability studies of morphine and rhodamine 123 demonstrated vectorial transport. The basolateral to apical (B:A) fluxes of morphine (50 microM) and rhodamine (1 microM) were approximately 50 and 100% higher than the fluxes from the apical to the basolateral direction (A:B), respectively. Decreasing the extracellular concentration of morphine to 0.1 microM resulted in a 120% difference between the B:A and A:B permeabilities. The addition of GF120918 abolished any significant directionality in transport rates across the endothelial cells. Efflux studies showed that the loss of morphine from BBCECs was temperature- and energy-dependent and was reduced in the presence of P-gp inhibitors. These observations indicate that morphine is transported by P-gp out of the brain capillary endothelium and that the BBB permeability of morphine may be altered in the presence of P-gp inhibitors.     

Cationic ligand appended nanoconstructs: a prospective strategy for brain targeting

The objective of present research was to evaluate the potential of engineered solid lipid nanoparticles (SLNs) as vectors to bypass the blood brain barrier. Anti-cancer agent, doxorubicin (DOX) loaded SLNs were prepared and conjugated with cationic bovine serum albumin (CBSA). The formation of CBSA tethered and plain SLNs were characterized by FTIR, NMR, and TEM analyses. Physicochemical parameters such as particle size/polydispersity index and zeta-potential were also determined. Cellular uptake studies on HNGC-1 cell lines depicted almost six times enhanced uptake of ligand conjugated SLNs as compared to plain DOX solution. Furthermore, CBSA conjugated formulation was more cytotoxic as compared to free drug or unconjugated SLNs. Transendothelial studies showed maximum transcytosis ability of CBSA conjugated SLNs across brain capillary endothelial cells. In vivo pharmacokinetic parameters and biodistribution pattern demonstrated efficiency of the system for spatial and temporal delivery of DOX to brain tissues. Lastly, hematological, nephrotoxic as well as hepatotoxic data suggested CBSA conjugated formulations to be less immunogenic compared to plain formulations.     

P-glycoprotein influences the brain concentrations of cetirizine (Zyrtec), a second-generation non-sedating antihistamine

Recent in vitro studies have suggested that P-glycoprotein (Pgp) and passive membrane permeability may influence the brain concentrations of non-sedating (second-generation) antihistamines. The purpose of this study was to determine the importance of Pgp-mediated efflux on the in vivo brain distribution of the non-sedating antihistamine cetirizine (Zyrtec), and the structurally related sedating (first-generation) antihistamine hydroxyzine (Atarax). In vitro MDR1-MDCKII monolayer efflux assays demonstrated that cetirizine was a Pgp substrate (B-->A/A-->B + GF120918 ratio = 5.47) with low/moderate passive permeability (PappB-->A = 56.5 nm/s). In vivo, the cetirizine brain-to-free plasma concentration ratios (0.367 to 4.30) were 2.3- to 8.7-fold higher in Pgp-deficient mice compared with wild-type mice. In contrast, hydroxyzine was not a Pgp substrate in vitro (B-->A/A-->B ratio = 0.86), had high passive permeability (PappB-->A + GF120918 = 296 nm/s), and had brain-to-free plasma concentration ratios >73 in both Pgp-deficient and wild-type mice. These studies demonstrate that Pgp-mediated efflux and passive permeability contribute to the low cetirizine brain concentrations in mice and that these properties account for the differences in the sedation side-effect profiles of cetirizine and hydroxyzine.     

Clinical pharmacokinetics of doxorubicin in combination with GF120918, a potent inhibitor of MDR1 P-glycoprotein

Previous clinical investigations with doxorubicin indicated that modulators of P-glycoprotein dramatically decrease the systemic clearance of the drug, which complicates the interpretation of toxicity and response data. In the present study, we examined the pharmacokinetics of doxorubicin and GF120918, a novel potent P-glycoprotein inhibitor, in cancer patients in a search for more selective modulation of multidrug resistance (MDR). Seven cohorts (46 patients) received sequential treatments with doxorubicin alone by a 5 min i.v. bolus (50-75 mg/m2), oral GF120918 alone (50 mg q.d.-400 mg b.i.d.), and the combination of doxorubicin and GF120918. Serial blood and urine samples were taken during both treatment courses and analyzed for doxorubicin and its metabolite doxorubicinol by a liquid chromatographic assay. The pharmacokinetic characteristics of doxorubicin in the presence or absence of GF120918 indicate a very minor overall effect of the modulator, except at the highest combined dose level (i.e. 75 mg/m2 plus 400 mg b.i.d.). A limited number of patients experienced significantly increased exposure to doxorubicinol upon combined treatment, which was associated with concomitantly higher plasma levels of GF120918. Sigmoidal maximum-effect models revealed significant correlations (p<0.02) between the area under the curve of doxorubicinol and the percent decrease in neutrophils and platelets. Sigmoidicity factors in the fitted Hill equation were similar between both treatment courses, suggesting no pharmacodynamic potentiation of doxorubicinol myelotoxicity by GF120918. Our data indicate that GF120918 at the tested doses of combination treatment achieves plasma concentrations that reverse MDR in experimental models and it lacks the significant kinetic interaction with doxorubicin observed previously with other modulators. Hence, it may be possible in future trials to assess the contribution of a potent inhibitor of P-glycoprotein activity to the toxicity and activity of doxorubicin with the knowledge that profound plasma pharmacokinetic interactions are unlikely.     

The impact of pharmacologic and genetic knockout of P-glycoprotein on nelfinavir levels in the brain and other tissues in mice

Insufficient concentrations of protease inhibitors such as nelfinavir may reduce the effectiveness of HIV dementia treatment. The efflux transporter mdr1 product P-glycoprotein (P-gp) has been demonstrated to play a role in limiting nelfinavir brain levels. The goal of this study was to compare the effect of GF120918 (10 mg/kg, IV), a P-gp inhibitor, on intravenous nelfinavir (10 mg/kg) in vivo disposition and tissue penetration in P-gp-competent mdr1a/1b (+/+) mice versus P-gp double knockout mdr1a/1b (-/-) mice. Intravenous administration with the P-gp inhibitor GF120918 to mdr1a/1b (+/+) mice increased nelfinavir concentrations over a range of 2.3- to 27-fold, whereas nelfinavir distribution in mdr1a/1b (-/-) mice was 2- to 16-fold higher than that in their wild counterparts. Nelfinavir levels after GF120918 coadministration were higher in the heart, liver, and kidneys than those detected with mdr1a/1b knockout mice. In contrast, mdr1a/1b knockout mice exhibited higher nelfinavir levels in the brain (16.1-fold vs. 8.9-fold increase) and spleen (4.1-fold vs. 2.3-fold increase) compared to pharmacological inhibition with GF120918 in wild mice. Most notably, GF120918 provided tissue-specific effects in mdr1a/1b knockout mice with enhanced (p < 0.05) drug accumulation in the brain ( approximately 21-fold) and heart (3.3-fold). Our results suggest mdr1a/1b-independant mechanisms may also contribute to nelfinavir tissue distribution in mice.     

Design,characterization and in vitro evaluation of SMEDDS containing an anticancer peptide,linear LyP-1

Abstract

LyP-1 (CGNKRTRGC) is a nine amino acid peptide that shows high specificity for tumor lymphatics. The aim of this study was to develop lipid-based formulations containing the linear form of LyP-1 for lymphatic targeting. Self-microemulsifying drug delivery systems (SMEDDS) were designed for the delivery of linear LyP-1 by itself and as a solid dispersion (SD). Formulations were characterized in terms of physical stability, pH, morphological properties, droplet size distribution and zeta potential. Thermodynamically stable microemulsions were obtained with an average droplet size around 20?nm and zeta potential near neutrality. Cytotoxicity studies of blank and peptide-containing SMEDDS were carried out on Caco-2 cell line. Bioactivity studies of peptide-containing formulations were carried out on MDA-MB-231 breast cancer cell line. It was shown that blank and peptide-containing SMEDDS formulations were not cytotoxic to Caco-2 cell line. However, formulations containing the peptide and peptide SD led to a significant decrease in cell viability on breast cancer cells. It could be concluded that the SMEDDS formulations containing the linear LyP-1 were successfully developed for the lymphatic targeting of solid tumors in vitro.     

Predicting P-Glycoprotein Effects on Oral Absorption: Correlation of Transport in Caco-2 with Drug Pharmacokinetics in Wild-Type and mdr1a(-/-) Mice in Vivo

PURPOSE: Cell-based permeability screens are widely used to identify drug-P-glycoprotein (PGP) interaction in vitro. However, their reliability in predicting the impact of PGP on human drug pharmacokinetics is poorly defined. The aim was to determine whether a quantitative relationship exists between PGP-mediated alterations in Caco-2 permeability and oral pharmacokinetics in mice. METHODS: Two indicators of drug efflux were measured in Caco-2 for a group of 10 compounds, the ratio of A-B and B-A transport (R9B-A/A-B)) and the ratio of A-B transport in the presence and absence of a PGP inhibitor, GF120918 (R(GF)). These data were correlated with ratios of oral plasma levels in either mdr1a(-/-) or mdr1a/1b(-/-) and wild-type mice (R(KO/WT in vivo)) calculated from literature data on these compounds. RESULTS: A significant, positive correlation (r2 = 0.8, p < 0.01) was observed between RGF and R(KO/WT in vivo). In contrast, R(B-A/A-B), a more commonly used in vitro measure, showed a much weaker correlation with in vivo data (r2 = 0.33, p = 0.11). A strong correlation with R(GF) was also observed after correction of in vivo data for PGP effects on IV clearance. CONCLUSION: The increase in A-B drug permeability following inhibition of PGP in Caco-2 allows a reasonable prediction of the likely in vivo impact that PGP will have on plasma drug levels after oral administration.