REVIEW Lipid Formulation Strategies for Enhancing Intestinal Transport and Absorption of P-Glycoprotein (P-gp) Substrate Drugs: In vitro/In

vivo Case Studies

PANAYIOTIS P. CONSTANTINIDES,1 KISHOR M. WASAN2
1 2

Biopharmaceutical & Drug Delivery Consulting, LLC, Gurnee, IL 60031 Division of Pharmaceutics and Biopharmaceutics, University of British Columbia, Vancouver, Canada V6T123

Received 23 May 2006; revised 23 August 2006; accepted 24 August 2006 Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jps.20780

ABSTRACT: The intestinal efux pump, P-glycoprotein (P-gp), located in the apical membranes of intestinal absorptive cells, can reduce the bioavailability of a wide range of drugs which are substrates for this membrane transporter. In addition to anticancer and anti-HIV drugs, NCEs for other disease indications are P-gp substrates and there is considerable interest in inhibiting P-gp and thus increasing the bioavailability of these molecules. In this review article, an overview of P-gp and its role in drug transport and absorption will be presented rst and then formulation strategies to effectively inhibit P-gp will be discussed and compared. These strategies independently and in combination, are: (a) coadministration of another P-gp substrate/specic inhibitor, and (b) incorporation of a nonspecic lipid and/or polymer excipient in the formulation. The rst approach, although very effective in inhibiting P-gp, utilizes a second active compound in the formulation and thus imposes regulatory constraints and long development timelines on such combination products. Excipient inhibitors appear to have minimal nonspecic pharmacological activity and thus potential side effects of specic active compound inhibitors can be avoided. Case studies will be presented where specic active compounds, surfactants, polymers, and formulations incorporating these molecules are shown to signicantly improve the intestinal absorption of poorly soluble and absorbed drugs as a result of P-gp inhibition and enhanced drug transport in vitro. 2006 Wiley-Liss,
Inc. and the American Pharmacists Association J Pharm Sci 96:235248, 2007

Keywords: biopharmaceutics; drug transport; P-glycoprotein inhibition; active compounds; surfactants; lipid formulations; self-emulsifying drug delivery systems; poorly soluble drugs; case studies

OVERVIEW OF P-GLYCOPROTEIN AND ITS ROLE IN DRUG TRANSPORT AND ABSORPTION

P-glycoprotein (P-gp) is a membrane-bound transporter that mediates active transport, efux, of
Correspondence to: Panayiotis P. Constantinides (Tel: 847599-9496; Fax: 847-599-9496; E-mail: constantinpp@aol.com)
Journal of Pharmaceutical Sciences, Vol. 96, 235248 (2007) 2006 Wiley-Liss, Inc. and the American Pharmacists Association

a wide range of structurally unrelated drugs and other xenobiotics out of the cells.1 It is expressed along the entire length of the gut and also in the liver (canalicular membrane), kidney, blood-brain barrier, and placenta.1,2 Intestinal P-gp is located on the apical membranes of the epithelial cells. Utilizing the energy that is generated by hydrolysis of ATP,3 P-gp drives the efux of various substrates against a concentration gradient and thus reduces their intracellular concentration and in the case of drugs their oral bioavailability
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Figure 1. (A) Proposed models of P-gp action: (a) Pore model: drugs associate with P-gp in the cytosolic compartment and are transported out through a protein channel. (b) Flippase model: P-gp acts as a drug exporter by ipping drugs from the inner leaet of the plasma membrane to the outer leaet against a concentration gradient. (c) Vacuum Cleaner Model: intramembranous molecules that do not belong to the membrane, are recognized by P-gp, enter P-gp from the membranous site and leave the cell. (B) Schematic representation of the structure of P-gp (adapted with permission from the publisher, Ref. 3).

(Fig. 1A and B). The proposed models of P-gp action are shown in Figure 1A. In the Pore model (a), drugs associate with P-gp in the cytosolic compartment and are transported out through a protein channel. A main feature of the Flippase model (b) is that P-gp acts as a drug exporter by ipping drugs from the inner leaet of the plasma membrane to the outer leaet against a concentration gradient. Finally, in regard to Vacuum Cleaner Model (c), intramembranous molecules which do not belong to the membrane, are recognized by P-gp, enter P-gp from the membranous site and leave the cell. Amongst the proposed three mechanistic models, it appears that the Flippase model is the most widely accepted. A schematic representation of the structure of P-gp
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with the membrane spanning, endo- and exodomains, is shown in Figure 1B.3 This complex glycoprotein (MW of 170 kDa; Fig. 1A) is the product of the MDR1 gene and efuxes drugs without modication thereby conferring multidrug resistance. Approximately 50% of the anticancer drugs used clinically today are substrates for P-gp and include anthracyclines, vinca alkaloids, taxanes, epidophyllotoxins, mitomycins, camptothecins, and other.46 Given its broad specicity for anticancer drugs, overexpression of P-gp in many types of tumors appears to be prevalent and may be induced rapidly in response to chemotherapy. The link of P-gp inhibition to drug transport and absorption is shown in Figure 2 where the
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are other advantages and disadvantages in the use of the Caco-2 versus the MDCK model in reference to their culturing times and transepithelial electrical resistance (TEER) with the former cells exhibiting longer culturing times (23 weeks) and forming very tight junctions with higher TEER values. However, it should be kept in mind that any cell culture model may behave differently from the in vivo situation particularly due to the lack of gut content ow and basolateral blood ow.7 Thus, it not surprising that when the permeability of various compounds was plotted against lipophilicity, the resulting curve was found to be sigmoidal with a very sharp slope and this was correlated to variability in the actual drug absorption in vivo.7 Regardless of their limitations, cellular models continue to be a valuable tool to study drug transport and absorption of multiple compounds and formulations in a timely fashion.
Figure 2. P-gp inhibition, drug transport, and absorption interrelationships. This diagram represents a simple way to emphasize the link between P-gp inhibition and drug transport and absorption. The common intersection area represents in vitro/in vivo correlations. Drug absorption is dened as prehepatic concentration of the drug.

P-GLYCOPROTEIN SUBSTRATES AND INHIBITORS: CLASSIFICATIONS AND COMPARISONS

In recent years, identifying and developing selective and potent P-gp inhibitors has been a major focus area in drug development particularly in treating drug-resistant tumors. P-gp inhibitors are noncytotoxic compounds and when combined with drugs that are efuxed by P-gp, their intracellular concentration is maintained and sensitivity to these therapeutics is restored.69 The P-gp inhibitors developed and used clinically to date can be divided into two major classes, specic and nonspecic (Tabs. 1 and 2). There are three generations of specic P-gp inhibitors (Tab. 1). The rst generation incorporates compounds which tend to be less potent and

intersected area in the Venn diagram represents in vitro/in vitro correlations. Caco-2 and MDCK cells express P-gp and are amongst the most widely used cellular models to study drug absorption.7 Since Caco-2 cells are derived from human colon carcinoma cells, P-gp expression and inhibition may closely resemble in vivo P-gp expression and inhibition; although such correlations have not yet strictly established. MDCK cells originate from dog kidney and thus there may be signicant differences in their substrate and inhibitor specicities from those of the human transporter.7 There
Table 1.

P-gp Inhibitors: Approved Actives and New Chemical Entities

1st Generation: Developed for other indications (less potent and not selective) Examples: Verapamil, Cyclosporine 2nd Generation: Developed initially to reduce toxicity, more selective than 1st generation due to reduced toxicity Examples: PSC 833 (CsA analog) without immunosuppressive effect Dexverapamil without the cardiac effect of verapamil 3rd Generation: Designed to be more potent and selective inhibitors Examples: VX-710 (biricodar) GF120918 (elacridar) LY335979 (zosuquidar.3HCl) OC144-093

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Table 2. P-gp Inhibitors: Lipidic and Polymeric Excipients (A) Surfactants C8/C10 glycerol and PEG esters: Cremophor, Solutol HS-15, Labrasol, Softigen 767, Aconnon E Sucrose esters: Sucrose monolaurate Polysorbates: Tween 80, Tween 20 Tocopherol esters: a-tocopheryl-PEG-1000-succinate (TPGS) (B) Polymers Pluronic block copolymers (poloxamers): poly-(ethyleneoxide)/poly-(propyleneoxide) block copolymers Amphiphilic diblock copolymers: Methoxypolyethylene glycol-block-polycaprolactone (MePEG-b-PCL)

not selective and have primarily been developed for other indications with unwanted side effects at levels necessary to inhibit P-gp clinically.8 These compounds include, the calcium channel blocker, verapamil, and the immunosuppressive drug, CsA. PSC 833 an analog of cyclosporine without the immunosuppressive effect and dexverapamil, a verapamil analog without its cardiac effect, belong to the second generation P-gp inhibitors which were developed primarily to reduce the toxicity of the parent compound/drug.8 Finally, the third generation inhibitors are characterized as being more potent and selective. The following third generation inhibitors are at least 10-fold more potent than the earlier compounds and inhibit P-gp in the 30100 nM range:8 VX-710 (biricodar), GF 120918 (elacridar), LY335979 (zosuquidar.HCl), XR9576 (tariquidar), and OC 144-093. Coadministration of specic pharmacologically active compounds can effectively enhance the oral bioavailability of P-gp substrate drugs. These compounds, however, have their own pharmacological activity and thus potential drugdrug interactions and enhanced side effects in such combination drug products need to be considered and properly addressed. In addition, active compound inhibitors may also interact with P-gp in other organs/tissues as it has been shown with the new generation P-gp inhibitors.8 Thus, due to anticipated product development challenges and regulatory constraints with dosage forms incorporating two active compounds, there is a need to identify other P-gp inhibitors which have minimal nonspecic pharmacological activity. Nonspecic inhibitors of P-gp include surfaceactive compounds, such as lipids/surfactants and/or polymers (Tab. 2). These agents exhibit low nonspecic pharmacological activity unlike the potentially more serious side effects of the active
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compound inhibitors. Several excipients which are present in pharmaceutical formulations can indirectly inhibit P-gp through effects on the lipid membrane and thus enhance the intestinal membrane permeability and oral absorption of the substrate drug.10 Surfactant inhibitors, primarily nonionic ones, include polyoxyethylated/ pegylated surfactants, such as, polyoxyl 35 castor oil (Cremophor), PEG-15-hydoxystearate (Solutol HS-15), medium-chain (C8/C10) glycerol and PEG esters (Labrasol, Softigen 767 and Acconon E), polysorbates such as Polysorbate 20 and 80, sucrose esters such as sucrose monolaurate, an a-tocopheryl polyethylene glycol-1000-succinate (TPGS). Amongst polymers, effective P-gp modulators are pluronic block copolymers which include poly(ethylene oxide)/poly-(propylene oxide) block copolymers and amphiphilic diblock copolymers, such as methoxypolyethylene glycol-block-polycaprolactone (MePEG-b-PCL).11 Structurally as discussed earlier, P-gp is a membrane-bound protein composed of 12 transmembrane domains and 2 cytoplasmic nucleotide binding domains. Based on the widely accepted Flippase model (Fig. 1A), P-gp translocates hydrophobic molecules/drugs from the inner to the outer leaet of the lipid bilayer upon a conformational change induced by ATP binding to a cytoplasmic binding domain (Fig. 1B). Thus, its activity is modulated by the physical state of the lipid bilayer where the protein resides as it has been demonstrated by several studies.4,6,12,13 Hydrophobic substrates are rst partitioned into the lipid bilayer and then by lateral diffusion interact with the P-gp binding domain within the lipid membrane.12,13 Molecules which can interact with the lipid bilayer, such as, surfactants and block copolymers can modulate the activity of P-gp indirectly or nonspecically. An alteration in the uidity of the lipid membrane environment of P-gp
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by surfactants was shown to modulate drug efux from MDR cells by reduction in the P-gp ATPase activity.13 The mechanisms by which amphiphilic pluronic block copolymers modulate P-gp activity include both membrane permeability changes through a reduction in membrane microviscosity, and cellular depletion of ATP levels.10 While P-gp inhibition by certain surfactants10,14 22 was observed at concentrations below or at their critical micelle concentration (CMC), for the MePEG-b-PCL copolymers modulation of the Pgp activity was observed at concentrations above their CMC with little or no activity below the CMC.11 It has been suggested that vesicular transport of micellized substrate may be one of the possible mechanisms by which these molecules modulate P-gp activity.11 Table 3 summarizes several biopharmaceutical effects of lipid-based systems on the absorption of the various classes of drug molecules,23 according to the Biopharmaceutics Classication Scheme.24 In addition to improving the solubilization of poorly soluble drugs, other biopharmaceutical effects of lipids include inhibition of enzymatic hydrolysis and drug efux. Due to these effects, intestinal absorption and oral bioavailability enhancement have been well established and representative case studies will be presented in subsequent sections of this Review article.

of the activity of P-gp using lipidic and polymeric excipients. The following case studies are representative of this rapidly growing eld of biopharmaceutics. Paclitaxel Proprietary lipid polymer emulsions (LPETM) incorporating P-gp inhibitors were developed and assessed for their ability to enhance paclitaxel transport in Caco-2 cells and to correlate this effect to intestinal drug absorption enhancement in rats.25,26 The ability of LPETM formulations to inhibit P-gp in Caco-2 cell monolayers was assessed using a calcein extrusion assay.27 Monolayers of Caco-2 cells were incubated with test formulations and controls and then with calceinAM, a uorescent P-gp substrate. After washing, the amount of intracellular calcein was measured with a uorescent plate reader. The data in Figure 3A demonstrated that both LPETM-S, incorporating C8/C10 glycerol and PEG esters (Softigen 767) and LPETM-CsA formulations were effective inhibitors of P-gp. Incorporation of CsA into LPETM did not affect its P-gp inhibitory activity. LPETM formulations incorporating Softigen 767 inhibited P-gp at 50%60% of the level of inhibition observed by CsA alone (positive control). Formulations containing a constant amount of paclitaxel were incubated with Caco-2 cells grown on Transwell lters. The transwells were removed after the transport period and Jurkat cells in the receiver wells were incubated for an additional 48 h. The viability of the Jurkat cells was then assessed by their ability to reduce the tetrazolium dye WST-I.28,29 Figure 3B shows the

P-gp INHIBITION BY LIPID EXCIPIENTS AND FORMULATIONS: IN VITRO CASE STUDIES

As discussed in the preceding section, there are many reports in the literature on the modulation
Table 3. Ref. 23)

Biopharmaceutical Effects of Oral Lipid-Based Systems (Modied from

BCS I II III

Aqueous Solubility High Low High

Membrane Permeability High High Low

Potential Effect of Lipid-Based Systems Enzymatic degradation # Gut wall efux # Solubilization " Bioavailability " Enzymatic degradation # Gut wall efux # Bioavailability " Solubilization " Enzymatic degradation # Gut wall efux # Bioavailability "

IV

Low

Low

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tions of TPGS. The apical-to-basolateral (A ! B) permeability of paclitaxel was found to increase in the presence of TPGS while the basolateral-toapical (B ! A) was decreased with maximum effect at 0.1 mg/mL of TPGS.30 Increasing TPGS concentrations above 0.1 mg/mL, resulted in a decrease in A ! B permeability with no change in the B ! A permeability, which suggests the involvement of monomeric and not micellar TPGS in P-gp inhibition.30 HIV Protease Inhibitors TPGS with a CMC of 0.2 mg/mL was also shown to improve the solubility and intestinal permeability of amprenavir, a protease inhibitor marketed as Agenerase1 by GSK.31 When the concentration of TPGS was increased above 0.1 mg/mL, an improvement in paclitaxel solubility was observed presumably via micellar solubilization similarly to the reported effect of TPGS on paclitaxel solubility.30 The effects of TPGS were reported to be: (a) improvement in the solubility (S) and thus dissolution of amprenavir through micellar solubilization, (b) enhancement of the permeability (Peff) of amprenavir across the gut wall, presumably through drug efux inhibition. In one particular study, TPGS enhanced the overall absorption ux (J Peff S) of the drug (Fig. 4) by increasing its solubility and intestinal permeability.31 To correlate the inhibitory effects of TPGS on drug efux to the structure of TPGS and specically to the chain-length of the PEG group, several

Figure 3. (A) Inhibition of P-glycoprotein by lipid excipient alone and in a lipid polymer emulsion (LPETM). S stands for Softigen 767, a C8/C10 mixture of glycerol and pegylated esters (B). Paclitaxel transport in Caco-2 cells in the presence of LPETM formulations incorporating P-gp inhibitors. S stands for Softigen 767, a C8/C10 mixture of glycerol and PEG esters.

concentrations of paclitaxel transported from LPETM-S and LPETM-CsA formulations and compared these values to those produced by media alone. Under the assay conditions there was no loss of monolayer integrity as determined by transepithelial resistance measurements data not shown). This data indicates that LPETM-CsA and LPETM-S formulations were effective in enhancing transport of paclitaxel across the Caco-2 monolayer although some transport was observed in the absence of these lipidic formulations. Enhancement of paclitaxel solubility and permeability by TPGS in vitro/in situ and effects on in vivo drug absorption has recently been reported by Varma and Panchagnula.30 In situ studies using rat ileum tissue in Ussing chambers, the bi-directional transport of 14C-paclitaxel was also monitored in the presence of increasing concentraJOURNAL OF PHARMACEUTICAL SCIENCES, VOL. 96, NO. 2, FEBRUARY 2007

Figure 4. Amprenavir ux in the presence of TPGS. Absorption ux levels off above 0.5 mg/mL of TPGS suggesting P-gp inhibition by monomeric TPGS (adapted from Ref. 31).
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analogs incorporating PEGs with an average MW in the range of 2006000 were synthesized and evaluated in vitro.32 The commercially available grade of TPGS contains PEG 1000 and its effect on the efux transport of the P-gp substrate Rhodamine 123 was compared against the other synthetic analogs in Caco-2 monolayers.32 The ability of these molecules to inhibit P-gp and enhance the apical to basolateral transport of Rhodamine 123 was also correlated to their cytotoxicity based on TEER measurements and lactate dehydrogenase (LDH) release. Interestingly, the results indicated an optimum PEG chain-length near 1000 which corresponds to the commercial TPGS product. Further mechanistic studies on these TPGS analogs are being pursued by the authors.32 Although TPGS is widely used as a drug solvent and oral bioavailability enhancer for poorly soluble drugs primarily by inhibiting P-gp, there have been cases where TPGS had a minimal or no effect on P-gp inhibition and drug absorption.19,3234 Differences in the physicochemical properties of the investigated compounds/drugs may account at least in part for the lack of a signicant P-gp inhibition in these studies. Celiprolol and Digoxin Studies reported by Cornaire et al.10 used the everted gut sac technique to study transport of two P-gp substrate drugs, Digoxin and Celiprolol. Digoxin (logP 1.51) undergoes intestinal and liver metabolism whereas celiprolol (logP 0.31) transport is stable in the intestine.10 The excipients were tested at 0.05, 0.1, and 0.5%, w/w and their effect on celiprolol tansport in the everted gut sac model is shown in Table 4.10 Cremophor EL and Acconon E had no effect whereas drug transport was enhanced by other surfactants in the order of Softigen 767 > TPGS > Imwitor 742.10 In the case of digoxin, the rank order was Labrasol > Imwitor 742 > Acconon E Softigen 767 > Cremophor EL > Miglyol > Solutol HS-15 > Sucrose Monolaurate > Polysorbate 20 > TPGS > Polysorbate 80.10 The relative effects of the investigated excipients appear to be dependent on the physicochemical properties of the P-gp substrate with differences in the lipophilicity of the P-gp substrate being a potential determining factor.10 Other properties such as the hydrophiliclipophilic balance (HLB) of the lipidic excipient may also be a contributing factor.10 It should also be mentioned that potential toxicity of the lipidic excipients should be considered in addressing
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Table 4. Effect of Surfactant Excipients on Celiprolol Absorption in the Everted Gut Sac (Adapted from Ref. 10) Excipient Concentration (%, w/w) Excipient Control Cremophor EL TPGS Acconon E Imwitor 742 Softigen 767 0.05 100% 91 58 52 11* 92 17 141 22 120 22 0.1 100% 87 68 53 15 50 12 123 20 96 13 0.5 100% 66 18 215 20** 96 34 189 24** 282 32**

Uptake is expressed as percentage of celiprolol uptake into the serosal contents of everted gut sacs in the presence of excipients compared with the control (no excipient). Reported values are means SE (n 3). *p < 0.05., the rest of the values are not signicant by comparison with the control. A two-way in reference to the time and concentrations of excipients ANOVA method with multiple comparisons (Fishers pairwise comparisons) was used for the statistical analysis of the data.10 **p < 0.01, the rest of the values are not signicant by comparison with the control. A two-way in reference to the time and concentrations of excipients ANOVA method with multiple comparisons (Fishers pairwise comparisons) was used for the statistical analysis of the data.10

their clinical use in various formulations. In the aforementioned studies, it was found that sucrose monolaurate and to a lesser extent Labrasol were toxic to the intestinal cells.10 Under the employed experimental conditions,10 these excipients produce 378% and 239% release of the cytosolic enzyme LDH under the experimental conditions used, respectively, relative to the 100% control value in the absence of any excipient.

Amphotericin B (AmpB) Amphotericin B (cLogP 1.29) is a heptaene macrolide antifungal derived from a strain of Streptomyces nodosus and used in the treatment of systemic fungal infections.35,36 AmpB is an amphoteric molecule with a carboxyl pKa of 5.7 and an amino pKa of 10. Due to its poor water solubility, AmpB is usually formulated as either a colloidal dispersion or in a lipid-based vehicle. Further studies were conducted by Wasan and colleagues to ascertain that the incorporation of AmpB into a glyceride-rich excipient Peceol (glyceryl monooleate) signicantly increased gastrointestinal absorption of AmpB in white male Sprague-Dawley rats.35,36 Based on preliminary studies,35 it was proposed that incorporation of AmpB into mixed micelles composed of Peceol
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would signicantly enhance gastro-intestinal (GI) tract absorption by decreasing P-glycoprotein (Pgp)-mediated drug efux. Caco-2 cells were seeded at 10000 cells/cm2 in T-75 asks. When the cells reached 80% conuency, they were treated for 1 day and 7 days with 0.1% to 1.0% (v/v) Peceol or media alone (control). Following treatment, total RNA was isolated using TRIzol1 reagent, followed by reverse transcription into single-stranded cDNA. Polymerase chain reactions (PCR) were performed with specic primers for mdr-1. The Pgp protein content was determined by Western Blot Analysis. A signicant lower mdr-1 mRNA and P-gp protein expression within Caco-2 cells was observed following 1- and 7-day treatment with Peceol 0.1% to 1.0% (v/v) compared to nontreated controls.36 Taken together, these ndings suggest that Peceol may increase the gastrointestinal absorption of AmpB by decreasing mdr1 mRNA and P-gp protein expression, resulting in lower PGP-mediated AmpB efux.36 Additional in vitro AmpB transport studies are warranted to further support this hypothesis.

EFFECTS OF LIPID FORMULATIONS INCORPORATING SPECIFIC AND NONSPECIFIC P-gp INHIBITORS ON DRUG TRANSPORT AND ABSORPTION: IN VIVO CASE STUDIES
There is increasing interest in the use of selfemulsifying drug delivery systems (SEDDS) to deliver orally poorly soluble drugs, including those which are substrates for P-gp.3739 Their compatibility with a variety of molecular structures of active compounds, oils, surfactants, and cosolvents and ease of processing and manufacture make SEDDS an attractive vehicle for pharmaceutical development. One advantage in using SEDDS with oral drugs that are P-gp substrates is their capacity to incorporate/solubilize high levels of specic and nonspecic P-gp inhibitors. This property will be illustrated in the subsequent case studies. In the case of TPGS and other tocols, advances in their use as drug delivery vehicles and specic in vitro/ in vivo case studies, including P-gp inhibition and oral absorption can also be found in a recently published review article.40 Cyclosporin A Absorption of CsA is affected by several factors such as poor solubility in aqueous solutions and
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GI uids, impaired bile ow and fat content in the diet.41 Sokol et al. presented clinical data obtained from liver transplantation patients, showing that TPGS enhances CsA absorption.42 Enhancement of the absorption of CsA was achieved when TPGS at 65 mg/kg/day was coadministered with the drug at doses in the range of 12.540 mg/kg/day compared to 29136 mg/kg/day in the absence of TPGS. Thus, coadministration of TPGS resulted in approximately 40%70% reduction in the daily dose of CsA. The role of TPGS in enhancing CsA absorption was further conrmed in clinical studies with liver transplantation patients.4345 A reduction in CsA daily dose and associated costs by TPGS without any clinical or biochemical evidence of TPGS induced toxicity, was reported by Pan et al.43 Boudreaux et al. reported on increased plasma concentration of CsA upon TPGS administration and this effect was attributed to enhanced drug solubilization through micelle formation.44 CsA pharmacokinetics in healthy subjects was affected by TPGS and a signicant increase in plasma AUC was reported.44 Independent studies have shown that the improvements in CsA pharmacokinetics were the result of the effects of TPGS in enhancing drug solubility and inhibiting P-gp and/or intestinal metabolism.46 Paclitaxel Paclitaxel, a highly hydrophobic and poorly water-soluble molecule and key chemotherapeutic agent with other disease indications in clinical development, is another oral drug with P-gp limited absorption. Reduction in the oral bioavailability of paclitaxel due to efux by P-gp has been demonstrated earlier by Sparreboom et al. in mice homozygous for a disruption of the mdr1a gene in comparison with normal mice.47 It was reported recently by Woo et al.48 that in the rat, approximately 54% of the dose of the orally administered paclitaxel is lost due to efux by P-gp, 38% due to gut lumen metabolism, and 3.5% by gut wall and liver metabolism. Improvement of the oral bioavailability of paclitaxel has been widely reported in the literature.25,26,4754 The most commonly used approaches were the coadministration of a specic P-gp substrate and/or incorporation of a nonspecic inhibitor in the formulation. Woo et al.48 using KR-30031, a verapamil analog with fewer cardiovascular effects than verapamil, showed that in Caco-2 cells the apical-to-basolateral transport of
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paclitaxel (5 mM) was increased in the presence of KR-30031 (25 mM) and this effect on drug efux was similar to that of verapamil (25 mM). A microemulsion formulation containing dimethylisosorbide, Tween 80 and DL-a-tocopherol and incorporating 6 mg/mL paclitaxel was administered by oral gavage to anesthetized rats.48 A dose of 25 mg/kg of paclitaxel was used with and without 5,10, 20, and 30 mg/kg of KR-30031 or 20 mg/kg of Verapamil. KR-30031 increased the oral bioavailability of paclitaxel from 4.6% to 41% (34% P-gp and 7% liver metabolism inhibition) in a dosedependent manner and this effect was saturable above 20 mg/kg of KR-30031.48 Earlier studies in improving the oral absorption of paclitaxel were focused on the coadministration of CsA or one of its analogs. By combining the marketed lipid formulations of paclitaxel (Taxol1) and CsA (Sandimmune1), the peroral bioavailability of paclitaxel in mice increased from 9.5% to 67%. Increase was 10-fold when CsA was substituted by its nonimmunosuppressive analog PSC833.54 In Phase II studies with cancer patients, weekly oral paclitaxel using the i.v.Taxol1 formulation, was administered in two doses of 90 mg/m2 on the same day proceeding with 10 mg/kg of CsA (Neoral1) given orally 30 min before.4951 It was found that oral paclitaxel was safe and active with plasma concentrations maintained for at least 150 ng/mL over 4 h. The Cmax and AUC did not increase as the absolute oral dose of paclitaxel was increased from 180 to 540 mg indicating a saturable process. The use of self-microemulsifying drug delivery system (SMEDDS) based on a-tocopherol and TPGS for enhancing the oral bioavailability of paclitaxel in rats has been reported by Yang et al.52

These SMEDDS are composed of a-tocopherol, TPGS, propylene glycol, sodium deoxycholate (DOC-Na), and Cremophor RH40. TPGS and DOC-Na in SMEDDS showed modest differences in the PK parameters of paclitaxel in rats (Table 5). However, when other specic P-gp inhibitors were included in the formulation, particularly CsA, signicant effects in these PK parameters were observed (Tab. 5). Thus, in these particular studies, signicant improvement in paclitaxel absorption was observed only upon a combination of lipidic excipient(s) with specic P-gp inhibiting drug. This in fact reects the actual clinical formulations of both Paclitaxel and CsA or another P-gp drug inhibitor, where P-gp inhibiting lipidic excipients are present in the formulations in order to solubilize and effectively deliver the drugs. Gao et al.53 developed a supersaturable selfemulsifying drug delivery system (S-SEDDS) for paclitaxel and other lipophilic drugs by incorporating hydroxypropyl methylcellulose (HPMC) in the formulation. By prolonging the supersaturated state, the presence of HPMC prevents drug precipitation that was observed from regular SEDDS formulations upon dilution with aqueous media to form a ne emulsion/microemulsion. The pharmacokinetics of paclitaxel from SEDDS and S-SEDDS was compared in rats upon oral administration of a paclitaxel dose of 10 mg/kg.53 A 10fold higher Cmax and a vefold higher AUC was obtained from S-SEDDS compared to SEDDS or the marketed Taxol1 formulation (9.5% absolute bioavailability from S-SEDDS vs. 2% from Taxol1 and 1% from SEDDS). In order to see further enhancement in the oral bioavailability of paclitaxel it was necessary to include cyclosporine in the formulation. In the presence of CsA (5 mg/kg)

Table 5. Pharmacokinetic Parameters of Paclitaxel after Oral Administration in Rats with P-gp Inhibitors (Modied from Ref. 52) 10.0 mg Paclitaxel/kg

2.0 mg Paclitaxel/kg SMEDDS (0.5% w/w) SMEDDS (0.5% w/w) SMEDDS (0.5% w/w)

5.0 mg Paclitaxel/kg

Taxol1 SMEDDS SMEDDS SMEDDS (0.6% w/v) (1.25% w/w) (1.25% w/w) (2.5% w/w)

Parameters Tmax (h) Cmax (ng/mL) AUC024 (ng h/mL) Fr0 ! 24 (%)

CsA Etoposide Tacrolimus CsA CsA Tacrolimus CsA (40.0 mg/kg) (20.0 mg/kg) (8.0 mg/kg) (40.0 mg/kg) (40.0 mg/kg) (8.0 mg/kg) (40.0 mg/kg) 1.0 164 23 1135.5 188.8 1.0 84 10.2 837.5 139.2 1.0 58 9 838.0 139.3 2.0 175 25 1134.0 168.2 4.0 225 27 1698.5 252.0 1.0 67 12 921.0 136.6 4.0 239 24 1746.0 250.5

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Table 6. Bioavailability of Paclitaxel in Rats % Fa (Mean SD, n 3) 100 11.8 3.3 14.3 7.3 19.8 12.2

Formulation Taxol1 i.v. CsA (Neoral1) Taxol1 p.o.b LPETM-S, i.d.c LPETM-CsA, i.d.
a b c

% F (AUCi.d./p.o./AUCi.v.) (Doseiv/Dosei.d./p.o.) 100. Peroral. Intraduodenal.

the Cmax and AUC of paclitaxel although the effect of TPGS on both parameters was greater. The bioavailability of paclitaxel increased 6.3-fold and 4.2-fold with TPGS or Verapamil, respectively.30 Mechanistically, however, further work is needed to support whether TPGS enhances the oral biovailability of paclitaxel by improving drug dissolution through micellar solubilization and/or by other mechanisms. As discussed in the previous section, the presence of TPGS micelles diminishes P-gp inhibition in vitro.30,31 HIV Protease Inhibitors A soft gelatin formulation using a SEDDS and incorporating TPGS, PEG 400, and propylene glycol was used to increase the bioavailability of amprenavir.31 The softgel formulation containing 20% TPGS produced 69 8% absolute bioavailability in beagle dogs at a drug dose of 25 mg/kg. When the concentration of TPGS in the formulation was increased from 20% to 50% the absolute bioavailability was increased to 80 16%.31 Although the difference in the bioavailability of amprenavir between softgel formulations incorporating 20% TPGS versus higher levels is small, the overall bioavailibility data in dogs were found to be correlated well with the in vitro Caco-2 permeability data as discussed earlier. It should also be pointed out that the 20%50% w/v TPGS or 200500 mg/mL is outside of the concentration range used in the study shown in Figure 4.31 However, this is in general the case with other studies, where the concentrations of surfactants used in in vivo absorption studies are much higher than those used in in vitro drug transport and permeability studies. Saquinavir, is a poorly soluble HIV protease inhibitor and it is marketed as Fortovase1 by Roche in a softgel formulation, 200 mg strength, where the drug is solubilized in a mixture of vitamin E and medium-chain mono- and diglycerides.55 A threefold increase in the oral bioavailability of saquinavir from the softgel formulation was reported compared to the solid dosage form Invarase1. With the reported oral bioavailability of saquinavir in Invarase1 being variable (1%9%) and averaged at about 4%, the drug bioavailability from the Fortovase1 formulation which is not reported can be estimated to be about 15%.56 Celiprolol and Digoxin Absorption studies of digoxin and celiprolol were reported by Cornaire et al.10 using the rat model
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in the S-SEDDS, the oral bioavailability of paclitaxel increased to about 23%.53 Table 6 summarizes bioavailability data of paclitaxel upon peroral (p.o.) or intraduodenal (i.d.) administration to rats of LPETM formulations incorporating P-gp inhibitors.25,26 The absolute bioavailabilities (% F) of paclitaxel in rats upon i.d. administration of LPETM-CsA and LPETM-S formulations were in the range of 15%20% using i.v. Taxol1 as a reference formulation, although with large interanimal variability (Tab. 6). The data suggest that LPETM formulations with and without CsA were at least as effective in improving the intestinal absorption of paclitaxel in rats as the combination of the marketed oral and i.v. formulations of these drugs, respectively. The peroral bioavailability of paclitaxel from the i.v. Taxol1 formulation was not investigated in these studies but it is expected to have a mean value of about 4%.30,47 51 The effects of the LPETM to inhibit P-gp activity and to enhance transport of paclitaxel across Caco-2 cells (Figs. 3A and B) correlated fairly well with the in vivo absorption of paclitaxel. These effects were more discriminatory in the P-gp inhibition data (Fig. 3A) than in drug transport data (Fig. 3B) possibly due to certain limitations of the employed drug transport system and assay. In the absence of additional studies, it is difcult to make absolute in vitro/in vivo correlations. One advantage of the LPETM drug delivery system is its ability to inhibit P-gp and improve drug transport and absorption in the absence of a second drug. As such, drug side effects and drugdrug interactions can be reduced. LPETM can be applied to other poorly absorbed water-insoluble drugs with P-gp limited intestinal absorption. In a recent study,30 14C-paclitaxel in a solution containing Cremophor: ethanol (1:1) that was further diluted with saline was administered perorally to anesthetized rats with and without the coadministration of TPGS (50 mg) or Verapamil (25 mg). Both TPGS and Verapamil increased
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and administering the formulations by oral gavage. Digoxin, dosed at 0.25 mg/kg, was dissolved at 0.025% (w/v) in water, propylene glycol, and ethanol (50:40:10, v/v/v) whereas celiprolol dosed at 30 mg/kg, was dissolved in water to produce a 1% w/v solution.10 The coadministered excipients were added to the drug solutions at the concentrations of 1 and 120 mg/kg for digoxin and celiprolol, respectively, to give the same drug:excipient ratio for each.10 The evaluated excipients changed the PK prole of the orally administered digoxin or celiprolol but without changing the overall AUC.10 Interestingly, only Softigen 767 produced a major increase in the AUC for the total absorption time course (0480 min). They observed an early peak of absorption (140 min), in the presence of certain excipients (TPGS, Imwitor 742, and Softigen 767) and interpreted it as being the result of higher excipient concentration in the proximal intestine where P-gp expression is lower.10 In the presence of these excipients, the AUC for the time course of 040 min for digoxin absorption was increased from 252.2 mg min/L of the control value to about 440 mg min/L.10 It is apparent from these and other results that the effects of lipid excipients/ surfactants on efux transport and the pharmacokinetic parameters of the administered drug are complex with multiple mechanisms being involved. The excipients are also mixtures of various chemical species with varying degree of purity and lot-to-lot variability and these characteristics should also be considered in interpret-

ing the results from in vitro and in vivo studies and ranking the excipients in terms of their relative effects. Amphotericin B Wasan and colleagues recently reported on the oral absorption of the polyene macrolide antifungal agent, AmpB, when incorporated into Peceol.35 The purpose of this study was to determine the effects of various lipid and mixed-micelle formulations on the oral absorption and renal toxicity of AmpB in rats. The maximum concentration of AmpB in plasma and the AUC024 h for AmpB were elevated in rats administered glyceride-rich AmpB formulations in comparison to those in rats given (i) AmpB preformulated as a micelle containing sodium deoxycholate with sodium phosphate as a buffer (DOC-AmpB), (ii) an AmpB-lipid complex suspension, or (iii) AmpB solubilized in methanol (Table 7). Furthermore, these ndings suggest that AmpB incorporated into glyceride-based oral formulations has less renal toxicity than DOC-AmpB.

CONCLUSIONS AND FUTURE PERSPECTIVES

Lipid excipients and formulations incorporating these excipients and/or specic active compounds continue to serve as a useful approach to inhibit P-gp and improve the oral bioavailability of drugs

Table 7. Plasma Creatinine and Amphotericin B (AmpB) Concentrations after Administration of a Single Intravenous Dose of Deoxycholate (DOC)-AmpB and a Single Oral Gavage of DOC-AmpB and Various AmpB Lipid Formulations to Male Sprague-Dawley Rats (Modied from Ref. 35) Plasma Creatinine Level Dose (mg/kg) 1 5 50 50 50 50 5 Prior to dose (mg/dL) 0.38 0.13 0.34 0.07 0.29 0.07 0.45 0.01 0.27 0.02 0.48 0.10 0.44 0.06 AUC024 h (ng h/mL) 4.3 1.1 ND 519 209** 542 271 5984 3461*** 11407 4971*** 4415 2411***

Treatment Groups

24 h 0.71 0.10* 0.29 0.05 0.42 0.02* 0.24 0.07* 0.38 0.02* 0.65 0.07 0.39 0.06

Tmax (h) ND 8 10 2 4 2

Cmax (ng/mL) ND 39.8 22 48.5 26 769 213*** 1469 891*** 1187 409***

DOC-AmpB IV Oral Oral AmpB lipid formulations ABLC Intralipid-AmpB Peceol/AmpB Peceol/AmpB

Data presented as mean standard deviation n 6 weighted 350400 g. *p < 0.05 versus prior to dose. **p < 0.05 versus IV DOC-AmpB. ***p < 0.05 versus AUC024 h for ABLC.
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with P-gp limited absorption. Based on the data reported in this review article, there appears to be a reasonably good correlation between the in vitro effects of lipidic excipients on P-gp inhibition and drug transport and in vivo drug absorption. In vitro/in vivo correlations, however, are not always clear and applicable to all classes of drugs with P-gp limited absorption. In cases where in vtro/in vivo correlations are established, there are certain product development and clinical requirements that also need to be considered such as: (a) the molecule must be specic and pharmacokinetic interactions must be minimal to reduce potential toxicity and (b) clinically relevant doses of the inhibitor and optimum duration of P-gp inhibition need to be considered. There are additional needs in the case of excipient inhibitors which include better understanding of the inhibition mechanism(s), and establishment of excipient inhibitor-P-gp substrate structureactivity relationships. To this end, the development of new and safe excipients with a wide spectrum of physicochemical and biopharmaceutical properties should be very benecial. It is evident that the function of excipients present in pharmaceutical formulations goes well beyond their use as inactive components to solubilize and stabilize drug molecules. Their expanding role as P-gp modulators requires that their function in the formulation is properly reassessed as more in vitro/in vivo data emerge using a wide range of excipients and P-gp substrates. Product development challenges, regulatory quality and acceptance of formulations incorporating these excipients can be assured once the above requirements are met.

B!A CsA TPGS GSK AmpB GI SEDDS AUC AUC0 ! 24 Cmax Tmax Fr SMEDDS DOC-Na S-SEDDS HPMC HIV PK

basolateral to apical transport cyclosporin A a-tocopheryl-PEG-1000-succinate GlaxoSmithKline amphotericin B gastro-intestinal self-emulsifying drug delivery system area-under-the-curve area-under-the-curve from 0 to 24 h maximum plasma concentration time to reach peak plasma concentration (Cmax) relative bioavailability self-microemulsifying drug delivery system sodium deoxycholate supersaturable self-emulsifying drug delivery system hydroxypropyl methylcellulose human immunodeciency virus pharmacokinetic

REFERENCES
1. Ueda K, Yoshida A, Amachi T. 1999. Recent progress in P-glycoprotein research. Anticancer Drug Design 14:115121. 2. Hunter J, Hirst BH. 1997. Intestinal secretion of drugs. The role of P-glycoprotein and related drug efux systems in limiting oral drug absorption. Adv Drug Del Rev 25:129157. 3. Varma MV, Ashokraj Y, Dey CS, Panchagnula R. 2003. P-glycoprotein inhibitors and their screening: A perspective from bioavailability enhancement. Pharmacol Res 48:347359. 4. Romsicki Y, Sharon FJ. 1999. The membrane lipid environment modulates drug interactions with the P-glycoprotein multidrug transporter. Biochemistry 38:68876896. 5. Higgins CF, Gottesman MM. 1992. Is the multidrug transporter a ippase? Trends Biochem Sci 17:1821. 6. Eytan GD, Regev R, Oren G, Assaraf YG. 1996. The role of passive transbilayer drug movement in multidrug resistance and its modulation. J Biol Chem 271:1289712902. 7. Pelkonen O, Boobis AR, Gundert-Remy U. 2001. In vitro prediction of gastrointestinal absorption and bioavailability: An experts meeting report. Eur J Clin Pharmacol 57:621629. 8. Dantzig AH, De Alwis DP, Burgess M. 2003. Considerations in the design and development of transport inhibitors as adjuncts to drug therapy. Adv Drug Del Rev 55:133150.
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ABBREVIATIONS
P-gp ATP MW MDR Caco-2 MDCK TEER NCE PEG CMC LPETM Calcein-AM A!B P-glycoprotein adenosine triphosphate molecular weight multi-drug resistance human colon adenocarcinoma cells Madin-Darby canine kidney cells transepithelial electrical resistance new chemical entity polyethylene glycol critical micelle concentration lipid polymer emulsions 6-carboxyuoresceinacetoxymethylester apical to basolateral transport

JOURNAL OF PHARMACEUTICAL SCIENCES, VOL. 96, NO. 2, FEBRUARY 2007

LIPID FORMULATION STRATEGIES FOR P-gp INHIBITION

247

9. Seelig A, Landwojtowicz E. 2000. Structure-activity relationship of P-glycoprotein substrates and modiers. Eur J Pharm Sci 12:3140. 10. Cornaire G, Woodley J, Hermann P, Cloarec A, Arellano A, Houin G. 2004. Impact of excipients on the absorption of P-glycoprotein substrates in vitro and in vivo. Int J Pharm 278:119131. 11. Zastre J, Jackson J, Burt H. 2004. Evidence for modulation of P-glycoprotein-mediated efux by methoxypolyethylene glycol-block-polycaprolactone amphiphilic diblock copolymers. Pharma Res 21:14891497. 12. Sinicrope FA, Dudeja PK, Bissonnette BM, Safa AR, Brasitus TA. 1992. Modulation of P-glycoprotein-mediated drug transport by alterations in lipid uidity of rat liver canalicular membrane vesicles. J Biol Chem 267:2499525002. 13. Regev R, Assaraf YG, Eytan GD. 1999. Membrane uidization by ether, other anesthetics, and certain agents abolishes P-glycoprotein ATPase activity and modulates efux from multi-drugresistant cells. Eur J Biochem 259:1824. 14. Bogman K, Erne-Brand F, Alsenz J, Drewe J. 2003. The role of surfactants in the reversal of active transport mediated by multidrug resistance proteins. J Pharm Sci 92:12501261. 15. Buckingham LE, Balasubramanian M, Emanuele RM, Clodfelter KE, Coon JS. 1995. Comparison of solutol HS15, cremophor EL and novel ethoxylated fatty acid surfactants as multidrug resistance modication agents. Int J Cancer 62:436442. 16. Hugger ED, Novak BL, Burton PS, Audus KL, Borchardt RT. 2002. A comparison of commonly used polyethoxylated pharmaceutical excipients on their ability to inhibit P-glycoprotein activity in vitro. J Pharm Sci 91:19912002. 17. Lo Y. 2003. Relationships between the hydrophiliclipophilic balance values of pharmaceutical excipients and their multidrug resistance modulating effect in Caco-2 cells and rat intestines. J Control Rel 90:3748. 18. Rege BD, Yu LX, Hussain AS, Polli JE. 2001. Effect of common excipients on Caco-2 transport of lowpermeability drugs. J Pharm Sci 90:17761786. 19. Rege BD, Kao JP, Polli JE. 2002. Effects of nonionic surfactants on membrane transporters in Caco-2 cell monolayers. Eur J Pharm Sci 16:237 246. 20. Anderberg EK, Nystrom C, Artursson P. 1992. Epithelial transport of drugs in cell culture. VII: Effects of pharmaceutical surfactant excipients and bile acids on transepithelial permeability in monolayers of human intestinal epithelial (Caco-2 cells). J Pharm Sci 81:879887. 21. Yamazaki T, Sato Y, Hanai M, Mochimaru J, Tsujino I, Sawada U, Horie T. 2000. Non-ionic detergent Tween 80 modulates VP-16 resistance in classical multidrug resistant K562 cells via

22.

23.

24.

25.

26.

27.

28.

29.

30.

31.

32.

33.

enhancement of VP-16 inux. Cancer Lett 149: 153161. Lo YL, Huang JD. 2000. Effects of sodium deoxycholate and sodium caprate on the transport of epirubicin in human intestinal epithelial Caco-2 cell layers and everted gut sacs of the rats. Biochem Pharmacol 59:665672. Constantinides PP. 1995. Lipid microemulsions for improving drug dissolution and oral absorption: Physical and biopharmaceutical aspects. Pharm Res 12:15611572. Amidon GL, Lennernas H, Shah V, Crison JR. 1995. A theoretical basis for a biopharmaceutic drug classication: The correlation of in vitro drug product dissolution and in vivo bioavailability. Pharm Res 12:413420. Fast DJ, Patil RT, Bosman KS, Loy LP, Constantinides PP. 2002. Enhancement of paclitaxel transport and intestinal absorption using lipid polymer emulsions (LPETM) incorporating Pglycoprotein inhibitors. International Symposium on Tumor Targeted Delivery Systems, Bethesda, MD. Fast DJ, Patil RT, Bosman KS, Constantinides PP. 2002. Lipid polymer emulsions (LPETM) incorporating P-glycoprotein inhibitors enhance the intestinal absorption of paclitaxel. Annual Meeting of the American Association of Pharmaceutical Scientists, Toronto, Canada. Tiberghien F, Loor F. 1996. Ranking of P-glycoprotein substrates and inhibitors by a calcein-AM uorometry screening assay. Anticancer Drug 7:568578. Artursson P, Karlsson J, Ocklind G, Shipper N. 1996. 1. Studying transport processes in absorptive epithelia. In: Shaw AJ, editor. Epithelial cell culture: A practical approach. New York: IRL Press. Rubas W, Cromwell MEM, Mrsny RJ, Ingle G, Elia KA. 1996. An integrated method to determine epithelial transport and bioactivity of oral drug candidates in vitro. Pharm Res 13:2326. Varma MVS, Panchagnula R. 2005. Enhanced oral paclitaxel absorption with vitamin E-TPGS: Effect of solubility and permeability in vitro, in situ and in vivo. Eur J Pharm Sci 25:445453. Yu L, Bridgers A, Polli J, Vickers A, Roy A, Winnike R, Cofn M. 1999. Vitamin E-TPGS increases absorption ux of an HIV Protease Inhibitor by enhancing its solubility and permeability. Pharm Res 16:18121817. Collnot E-V, Baldes C, Wempe MF, Hyatt J, Navarro L, Edgar KV, Schaefer UF, Lehr C-MJ. 2006. Inuence of vitamin E TPGS poly(ethylene glycol) chain length on apical efux transporters in Caco-2 cell monolayers. Control Rel 111:3540. Johnson BM, Charman WN, Porter CJ. 2002. An vitro examination of the impact of polyethylene glycol 400, Pluronic P85, and vitamin-E D-alphatocopheryl-polyethylene glycol 1000-succinate on P-glycoprotein efux and enterocyte-based

DOI 10.1002/jps

JOURNAL OF PHARMACEUTICAL SCIENCES, VOL. 96, NO. 2, FEBRUARY 2007

248

CONSTANTINIDES AND WASAN

34.

35.

36.

37. 38.

39.

40.

41.

42.

43.

44.

metabolism in excised rat intestine. AAPS Pharm Sci 4:E40. Ramsay-Olocco K, Alexandrova L, Nellore R, Killion R, Li L, Coen P, Ho Q, Jung D, Rocha C. 2004. Preclinical and clinical evaluation of solution and soft gelatin capsule formulations for BCS class 3 compound with atypical physicochemical properties. J Pharm Sci 93:22142221. Risovic V, Boyd M, Choo E, Wasan KM. 2003. Effect of various lipid-based oral formulations on plasma and tissue concentrations and renal toxicity of amphotericin B within male rats. Antimicrob Agents Chemother 47:33393342. Risovic V, Sachs-Barrable K, Boyd M, Wasan KM. 2004. Potential mechanisms by which peceol increases the gastrointestinal absorption of amphotericin B. Drug Dev Ind Pharm 30:767774. Pouton CW. 1997. Formulation of self-emulsifying drug delivery systems. Adv Drug Del Rev 25:4758. Gershanik T, Benita S. 2000. Self-dispersing lipid formulations for improving oral absorption of lipophilic drugs. Eur J Pharm Biopharm 50:179188. Constantinides PP. 2000. Self-emulsifying drug delivery formulations in the 21st century: Challenges and opportunities. In: Park K, Mrsny RJ, editors. Controlled drug delivery: Designing technologies for the future. ACS Symposium Series 752, Washington, DC: American Chemical Society, 284 296. Constantinides PP, Han J, Davis SS. 2006. Advances in the use of tocols as drug delivery vehicles. Pharm Res 23:243255. Odeberg JM, Kaufmann P, Kroon KG, Hoglund P. 2003. Lipid drug delivery and rational formulation design for lipophilic drugs with low oral bioavailability, applied to Cyclosporine. Eur J Pharm Sci 20:375382. Sokol RJ, Johnson KE, Karrer FM. 1991. Improvement of cyclosporine absorption in children after liver transplantation by means of water-soluble vitamin E. Lancet 338:211215. Pan SH, Lopez RR, Sher LS, Hoffman AL, Podesta LG, Makowka L, Rosenthal P. 1996. Enhanced oral cyclosporine absorption with water-soluble vitamin E early after liver transplantation. Pharmacother 16:5965. Boudreaux JP, Hayes DH, Mizrahi S, Maggiore P, Blazek J, Dick D. 1993. Use of water-soluble liquid vitamin E to enhance cyclosporine absorption in children after liver transplant. Transplant Proc 25:18751881.

45. Chang T, Benet LZ, Herbert MF. 1996. The effect of water-soluble vitamin E on cyclosporine pharmacokinetics in healthy volunteers. Clin Pharmacol Ther 59:297303. 46. Dintaman JM, Silverman JA. 1999. Inhibition of Pglycoprotein by D-alpha-tocopheryl polyethylene glycol succinate (TPGS). Pharm Res 16:15501556. 47. Sparreboom A, Asperen J, Mayer U, Schinkel AH, Smit JW, Meijer DKF, Borst P, Nooijen WJ, Beijnen JH, Tellingen O. 1997. Limited oral bioavailability and active epithelial excretion of Paclitaxel (Taxol) caused by P-glucoprotein in the intestine. Proc Natl Acad Sci USA 94:20312035. 48. Woo JS, Lee CH, Shim CK, Hwang S-J. 2003. Enhanced oral bioavailability of paclitaxel by coadministration of the P-glycoprotein inhibitor KR30031. Pharm Res 20:2430. 49. Malingre MM, Beinjen JH, Schellens JHM. 2001. Oral delivery of taxanes. Invest New Drugs 19:155162. 50. Van Zuylen L, Verweij J, Sparreboom A. 2001. Role of formulation vehicles in taxane pharmacology. Invest New Drugs 19:125141. 51. Schellens JHM, Malingre MM, Kruijtzer CMF, Bardelmeijer HA, Tellingen O, Van Schinkel AH, Beijnen JH. 2000. Modulation of oral bioavailability of anticancer drugs: From mouse to man. Eur J Pharm Sci 12:103110. 52. Yang S, Gursoy RN, Lambert G, Benita S. 2004. Enhanced oral absorption of paclitaxel in a novel self-microemulsifying drug delivery system with or without concomitant use of P-glycoprotein inhibitors. Pharm Res 21:261270. 53. Gao P, Rush BD, Pfund WP, Huang T, Bauer JM, Morozowich W, Kuo MS, Hageman MJ. 2003. Development of a supersaturable SEDDS (S-SEDDS) formulation of paclitaxel with improved oral bioavailability. J Pharm Sci 92:2386 2398. 54. van Asperen J, van Tellingen O, Sparreboom A, Schinkel AH, Borst P, Nooijen WJ, Beijnen JH. 1997. Enhanced oral bioavailability of paclitaxel in mice treated with the P-glycoprotein blocker SDZ PSC 833. Br J Cancer 76:11811183. 55. Alsenz J, Steffen H, Alex R. 1998. Active apical secretory efux of the HIV protease inhibitors saquinavir and ritonavir in Caco-2 cell monolayers, Pharm Res 15:423428. 56. Strickley RG. 2004. Solubilizing excipients in oral and injectable formulations. Pharm Res 21:201 230.

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