Pharmaceutical Research, Vol. 22, No. 3, March 2005 (© 2005)
`DOI: 10.1007/s11095-004-1871-1
`
`Research Paper
`
`Intravenous Hydrophobic Drug Delivery: A Porous Particle Formulation of
`Paclitaxel (AI-850)
`
`Julie A. Straub,1,3 Donald E. Chickering,1 Jonathan C. Lovely,1 Huimin Zhang,1 Bhavdeep Shah,1
`William R. Waud,2 and Howard Bernstein1
`
`Received September 28, 2004; accepted December 13, 2004
`
`Purpose. To develop a rapidly dissolving porous particle formulation of paclitaxel without Cremophor
`EL that is appropriate for quick intravenous administration.
`Methods. A rapidly dissolving porous particle formulation of paclitaxel (AI-850) was created using spray
`drying. AI-850 was compared to Taxol following intravenous administration in a rat pharmacokinetic
`study, a rat tissue distribution study, and a human xenograft mammary tumor (MDA-MB-435) model in
`nude mice.
`Results. The volume of distribution and clearance for paclitaxel following intravenous bolus adminis-
`tration of AI-850 were 7-fold and 4-fold greater, respectively, than following intravenous bolus admin-
`istration of Taxol. There were no significant differences between AI-850 and Taxol in tissue concen-
`trations and tissue area under the curve (AUC) for the tissues examined. Nude mice implanted with
`mammary tumors showed improved tolerance of AI-850, enabling higher administrable does of pacli-
`taxel, which resulted in improved efficacy as compared to Taxol administered at its maximum tolerated
`dose (MTD).
`Conclusions. The pharmacokinetic data indicate that paclitaxel in AI-850 has more rapid partitioning
`from the bloodstream into the tissue compartments than paclitaxel in Taxol. AI-850, administered as an
`intravenous injection, has been shown to have improved tolerance in rats and mice and improved
`efficacy in a tumor model in mice when compared to Taxol.
`
`KEY WORDS: drug delivery; hydrophobic drugs; microparticles; paclitaxel; spray drying.
`
`INTRODUCTION
`
`The poor aqueous solubility of hydrophobic drugs is a
`challenging formulation problem, particularly for intravenous
`delivery (1). It is estimated that 40% of new chemical entities
`have poor aqueous solubility (2). Poor aqueous solubility is,
`in particular, a common property of compounds identified
`using combinatorial chemistry and high-throughput screening
`(3). Thus, creating intravenous formulations of such com-
`pounds is of significant value. Paclitaxel, the active ingredient
`in the commercially available anticancer agent Taxol (pacli-
`taxel injection, Bristol-Myers Squibb Oncology, Princeton,
`NJ, USA), has low aqueous solubility (4). Taxol consists of
`paclitaxel dissolved in Cremophor EL (BASF, Ludwigshafen,
`Germany; polyoxyethylated castor oil) and ethanol. Side ef-
`fects observed with the intravenous administration of Taxol
`such as hypersensitivity reactions, nephrotoxicity, and neuro-
`toxicity have been attributed to Cremophor (5). Due to such
`reactions, patients must be premedicated with corticosteroids
`and antihistamines, and Taxol must be administered as a 1–24
`h infusion (6–7). Taxol is an important treatment of breast,
`ovarian, and non-small cell lung carcinomas (8–9), and modi-
`fied formulations of paclitaxel that do not contain Cremophor
`
`1 TAcusphere, Inc., Watertown, Massachusetts 02472, USA.
`2 Southern Research Institute, Birmingham, Alabama 35205, USA.
`3 To whom correspondence should be addressed. (e-mail: julie.
`straub@acusphere.com)
`
`are currently of significant interest (10–12). A number of
`nanoparticle- and microparticle-based formulations of pacli-
`taxel, and related formulations such as liposomes and emul-
`sions, have been reported in the literature (13–21). In these
`formulations, the paclitaxel is encapsulated within another
`material such as a hydrophobic polymer like poly (lactide
`co-glycolide) (PLGA), a protein, or lipids. These types of
`materials create a sustained-release system for paclitaxel with
`drug release occurring over hours to months. The porous par-
`ticle formulation of paclitaxel (AI-850) discussed in this paper
`was developed with the goal of creating a rapidly dissolving
`formulation that could be administered as an i.v. bolus or a
`short infusion.
`Acusphere’s hydrophobic drug delivery system (HDDS),
`consisting of sub-micrometer to micrometer size porous par-
`ticles, can be used to create rapidly dissolving formulations of
`hydrophobic drugs (22–25). The porous particles consist of
`drug microparticles and sub-micrometer particles and a wa-
`ter-soluble excipient such as a sugar or amino acid. During
`reconstitution, the water-soluble excipient dissolves, leaving a
`suspension of drug microparticles and sub-micrometer par-
`ticles that dissolve upon further dilution in the plasma. The
`porous nature of the particles facilitates wetting and rapid
`dissolution of the encapsulated drug.
`The production of the porous particles involves spray
`drying a solution containing the drug, the water-soluble ex-
`cipient, and a pore-forming agent (i.e., a volatile salt). The
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`Straub et al.
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`pore-forming agent is volatilized along with the process sol-
`vent during the spray drying to produce a porous matrix com-
`prising drug microparticles and the excipient. The water-
`soluble excipient facilitates wetting of the drug particles dur-
`ing reconstitution, provides proper osmolality to the dosage
`form, and improves the storage stability of the dry powder.
`The spray-drying process can be performed aseptically, and
`the resulting particles are in the appropriate size range for i.v.
`administration following reconstitution, as well as appropri-
`ate for administration by other parenteral routes, oral admin-
`istration, and pulmonary administration.
`This paper discusses the application of the hydrophobic
`drug delivery system to a rapidly dissolving formulation of
`paclitaxel for intravenous administration as an i.v. bolus or a
`short infusion and which would not require premedication of
`the patients with corticosteroids and antihistamines.
`
`MATERIALS AND METHODS
`
`Production of Microparticles
`
`A paclitaxel-containing solution was prepared by dissolv-
`ing paclitaxel (6.25 mg/ml), polysorbate 80 (0.625 mg/ml), and
`polyvinylpyrrolidone C15 (0.625 mg/ml) in ethanol. An aque-
`ous solution was prepared containing the pore-forming agent
`(22), ammonium bicarbonate, at 2.5 mg/ml, and mannitol at
`25 mg/ml. The aqueous solution was added to the ethanol
`solution in a ratio of 1:4. The resulting solution was spray
`dried on a spray dryer custom-designed to operate aseptically.
`The spray dryer was made from 316 stainless steel compo-
`nents with tri-clamp connections. The system was run under
`positive pressure with sterile-filtered nitrogen gas for the dry-
`ing gas and atomization gas. A 0.22-␮m filter was installed on
`the exhaust to maintain sterility and to capture any paclitaxel
`particulates that escape the spray-dryer cyclone.
`The spray dryer used an internal mixing air-atomizing
`nozzle (the internal diameter of the fluid cap was 0.028⬙, and
`the internal diameter of the air cap was 0.060⬙) and nitrogen
`as the drying gas. Spray-drying conditions were as follows: 75
`ml/min solution flow rate, 154 L/min atomization gas rate, 120
`kg/h drying gas rate, 116°C inlet temperature, and 62°C outlet
`temperature. The resulting powder was collected, filled into
`glass vials, and stored at 2–8°C.
`
`Paclitaxel Potency and Purity Analysis
`
`For potency and purity analysis, AI-850 was dissolved in
`methanol:water:acetic acid (85:15:1) and analyzed by HPLC
`using an Agilent 1100 Series high pressure liquid chromotog-
`raphy (HPLC) (Agilent Technologies Inc., Palo Alto, CA,
`USA). Chromatographic conditions used for the analysis of
`paclitaxel used UV detection at 230 nm and a Phenomenex
`Curosil PFP column (5 ␮m, 250 × 4.6 mm) held at 35°C during
`analysis. The mobile phases used were A (acetonitrile:water,
`35:65) and B (acetonitrile:water, 75:25), with a gradient run-
`ning from 100% A to 85% A over 12 min, followed by a
`gradient running from 85% A to 0% A over the next 11.5
`min. A flow rate of 1.5 ml/min was used. Potency values were
`obtained via comparison to a reference standard obtained
`from Hauser Laboratories (Boulder, CO, USA) and are
`based on 100% potency being the theoretical loading of the
`AI-850 powder based on the composition spray dried.
`
`Differential Scanning Calorimetry (DSC)
`
`DSC analyses were carried out on a TA 2920 differential
`scanning calorimeter (TA Instruments, New Castle, DE,
`USA) using nitrogen as the purge gas. Indium metal was used
`as the calibration standard. The samples were heated on the
`DSC at a heating rate of 10°C/min to a final temperature of
`350°C.
`
`Dry Powder Particle Size Analysis
`
`Particle size analysis of AI-850 powder (prior to recon-
`stitution) was performed using a Malvern Mastersizer (Mal-
`vern Instruments Ltd., Worcester, England) fitted with a dry
`powder module at a pressure of 65 psi.
`
`Particle Size Analysis Post-Reconstitution
`
`Particle size analysis of paclitaxel particles in AI-850
`post-reconstitution was performed using a Multisizer II
`(Beckman Coulter Inc., Fullerton, CA, USA) equipped with
`a 50-␮m aperture. The electrolyte used for the analysis con-
`tained sodium chloride (9 mg/ml), monobasic sodium phos-
`phate monohydrate (0.19 mg/ml), anhydrous dibasic sodium
`phosphate (1.95 mg/ml), mannitol (55 mg/ml), polyvinylpyrol-
`lidone K15 (5.5 mg/ml), and was adjusted to pH 7.4 with 0.1
`N HCl. The electrolyte solution was presaturated with pacli-
`taxel immediately prior to use by addition of a solution of
`paclitaxel in methanol (35 mg paclitaxel/ml methanol; 4 ml to
`1 L of modified electrolyte), followed by filtration through an
`0.22-␮m cellulose acetate filter to remove excess (precipi-
`tated) paclitaxel. The electrolyte was saturated with paclitaxel
`to ensure the paclitaxel particles in AI-850 did not dissolve
`upon dilution into the electrolyte for analysis. The presence
`of mannitol and polyvinylpyrollidone K15 in the electrolyte
`was found to suppress crystallization of paclitaxel from the
`paclitaxel-saturated electrolyte. The paclitaxel-saturated elec-
`trolyte was analyzed by Coulter Multisizer analysis prior to
`use for analysis of AI-850 suspensions to ensure that no sig-
`nificant crystallization of paclitaxel had occurred in the elec-
`trolyte solution. Suspensions of AI-850 were added to the
`paclitaxel-saturated electrolyte for analysis.
`
`Microscopy
`
`For transmission electron microscopy (TEM), AI-850
`powder was embedded in a LR White Resin, which was then
`cut into thin (100–120 nm) sections on a diamond knife, and
`the sections were imaged on a Zeiss EM-10 transmission elec-
`tron microscope (Zeiss Group, Jena, Germany) at 60 kV.
`For scanning electron microscopy (SEM), the particles
`were sputter coated with platinum-palladium (80:20) and then
`imaged on a Hitachi S-4800 or a Hitachi S-2700 (Hitachi High
`Technologies America, Inc., San Jose, CA, USA).
`For scanning electron microscopy (SEM) of paclitaxel
`particles post-reconstitution, the suspension post-recon-
`stitution was filtered using an 0.22-␮m filter, and the pacli-
`taxel particles retained on the filter were rinsed with water
`and vacuum dried prior to sputter coating.
`
`In Vitro Dissolution Analysis
`
`Dissolution studies were conducted in PBS containing
`0.08% Tween 80 (T80/PBS). T80/PBS (10 ml) was added to
`
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`Intravenous Hydrophobic Drug Delivery
`
`349
`
`an appropriate amount of material being tested to contain 5
`mg of paclitaxel in a 15 ml polypropylene conical tube, and
`the suspension was vortexed to create a suspension of micro-
`particles of paclitaxel. The suspension (0.25 ml) was then
`added to 250 ml of T80/PBS in a 600 ml glass beaker for
`dissolution analysis. Samples (1–2 ml) were removed and im-
`mediately filtered at each time-point. HPLC analysis was per-
`formed directly on the filtered aqueous solutions using an
`Agilent 1100 Series HPLC (Agilent Technologies Inc.). Chro-
`matographic conditions used for the analysis of dissolution of
`paclitaxel used a Nucleosil column (5 ␮m, C18, 100A, 250 ×
`4.6 mm), a mobile phase of 2 mM H3PO4/acetonitrile (2:3) at
`a flow rate of 1.5 ml/min, UV detection at 227 nm, and a run
`time of 25 min. The concentration of paclitaxel in the disso-
`lution media was 0.5 ␮g/ml. The saturation concentration ob-
`served for paclitaxel in T80/PBS was observed to be 1.5 ␮g/
`ml, and thus the dissolution assay was performed at a con-
`centration lower than the saturation concentration of
`paclitaxel in the dissolution media.
`
`Animal Care
`
`In the three animal studies described below, cage size
`and animal care conformed to the guidelines of the Guide for
`the Care and Use of Laboratory Animals, 7th Edition (Na-
`tional Research Council, National Academy Press, Washing-
`ton, DC, 1996), and the U.S. Department of Agriculture
`through the Animal Welfare Act (Public Law 99-198).
`
`Pharmacokinetic Study of Paclitaxel in Rats
`
`Male and female Sprague-Dawley rats (3/sex per group;
`animals were 8–11 weeks of age) were assigned to one of eight
`groups. Two groups received an i.v. bolus of Taxol (5 mg/kg
`or 10 mg/kg). Two groups received a 3 h i.v. infusion of Taxol
`(5 mg/kg or 10 mg/kg of paclitaxel). Four groups received an
`i.v. bolus of AI-850 (5 mg/kg, 10 mg/kg, 20 mg/kg, or 30 mg/kg
`of paclitaxel). AI-850 was reconstituted with water and di-
`luted with D5W for dosing at a paclitaxel concentration of 10
`mg/ml. Taxol was diluted with sterile saline for dosing at a
`paclitaxel concentration of 1 mg/ml.
`Serial blood samples (0.3 ml) were taken during 24 h and
`processed into plasma. For the bolus dose groups, blood
`samples were collected at 0 min (pre-dose), 5 and 30 min, and
`at 1, 2, 3, 5, 8, 12, and 24 h after dosing. For the infusion
`groups, blood samples were collected at 0 min (pre-infusion)
`and at the following time-points after the start of the infusion:
`30 min, and at 1, 2, 3, 4, 5, 8, 12, and 24 h. The plasma samples
`were analyzed for paclitaxel using a validated liquid chroma-
`tography/mass spectrometry/mass spectrometry (LC/MS/MS)
`assay at Southern Research Institute. The pharmacokinetic
`parameters were calculated assuming a two-compartment
`analysis using WinNonlin software.
`
`Tissue Distribution Study of Paclitaxel in Rats
`Single doses of either AI-850 or Taxol were administered
`intravenously via tail vein to groups of 18 male Sprague-
`Dawley rats/group. Animals were 8–9 weeks of age. The pac-
`litaxel dose was 5.7 mg/kg for both formulations. AI-850 was
`reconstituted with water and diluted into D5W to a paclitaxel
`concentration of 6.9 mg/ml for administration. Taxol Injection
`was diluted with saline to a final paclitaxel concentration of
`1.0 mg/ml for administration. Samples of plasma and selected
`
`tissues were collected from three rats/group at 0.5, 1, 2, 5, 12,
`and 24 h post-dose. The following tissues were collected and
`flash-frozen in liquid nitrogen: adrenal glands, brain, bone
`marrow, heart, kidney, liver, lung, muscle (back), small intes-
`tine (duodenum), sciatic nerve, spleen, testes, and thymus.
`Plasma and tissue samples were analyzed for concentrations
`of paclitaxel using a validated LC/MS/MS method (tissue
`samples were homogenized prior to analysis) at Southern Re-
`search Institute. Mean tissue paclitaxel concentration data
`were subjected to pharmacokinetic analyses using WinNonlin
`software. Values of AUC (area under the curve) were calcu-
`lated using the trapezoidal rule. Plasma paclitaxel concentra-
`tion data were fit to a compartmental model.
`
`In Vivo Antitumor Efficacy Study in Mice
`
`Mice were young (approximately 23 g), adult, female
`athymic NCr-Nu (14 mice/group). Mice were implanted sub-
`cutaneously with 30–40 mg fragments of the human mammary
`tumor MDA-MB-435 on day 0 (26). Treatment was scheduled
`to begin when the tumors ranged in size from 75 to 150 mg.
`Three groups were treated with AI-850 (15 mg/kg, 30 mg/kg,
`or 40 mg/kg of paclitaxel), one group was treated with the
`maximum tolerated dose (MTD) of Taxol (27) in this model
`(15 mg/kg of paclitaxel), and one control group was treated
`with D5W. AI-850 was reconstituted with 5% dextrose
`(D5W) to paclitaxel concentrations of 1.5 mg/ml, 3 mg/ml,
`and 4 mg/ml. Taxol was diluted with saline to a paclitaxel
`concentration of 1.5 mg/ml. Agents were administered via i.v.
`injection into the tail vein, once a day for 5 days. Mice were
`observed for survival, tumor size, and body weight. Tumors
`were measured by caliper in two dimensions and converted to
`tumor mass using the formula for a prolate elipsoid (l × w2/2).
`Antitumor activity was assessed by the delay in tumor
`growth of the treated groups in comparison to the vehicle-
`treated control. The values for the time required for two tu-
`mor mass doublings (four times the original size) and t-c were
`calculated. The value of t-c (days) is the difference in the
`median of times post-implant for tumors of the treated groups
`to attain an evaluation size compared to the median of the
`control group. The time required for tumor mass doubling is
`calculated based on the initial tumor weight at the beginning
`of the treatment period, and values between measurements
`are calculated by exponential extrapolation.
`
`RESULTS
`
`Analysis of Paclitaxel Within AI-850
`
`AI-850 was made via spray drying an ethanol-water so-
`lution containing paclitaxel, ammonium bicarbonate (the
`pore-forming agent), mannitol, polysorbate 80, and polyvinyl-
`pyrrolidone C15. As expected for a spray-drying process, pac-
`litaxel encapsulation efficiency was essentially 100%, with ob-
`served values of chromatographic potency of 99.8% ± 1.5%
`(n ⳱ 6 lots). The chromatographic purity of paclitaxel in
`AI-850 was comparable to the purity of the starting bulk pac-
`litaxel, indicating that the paclitaxel molecule was stable to
`the processing conditions. For example, the chromatographic
`purity of the paclitaxel for both the starting raw material and
`the lot of AI-850 used in the tissue distribution study were
`observed to be 99.5%. DSC analysis indicated that the pacli-
`taxel within AI-850 was amorphous.
`
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`Fig. 1. Dry powder particle size distribution of AI-850 as analyzed on
`the Malvern Mastersizer.
`
`Particle Characteristics
`
`The mean particle size for AI-850 prior to reconstitution
`was 1.53 ␮m ± 0.07 ␮m (n ⳱ 6 lots). A dry powder particle
`size distribution from a representative lot is shown in Fig. 1.
`The volume mean diameter is 1.52 ␮m, and 40% of the vol-
`ume is less than 1.0 ␮m. A scanning electron micrograph of
`AI-850 particles is shown in Fig. 2, in which the surface of the
`microparticles appears relatively smooth, and the micropar-
`ticles appear to be sphere-shaped. A transmission electron
`micrograph of cross-sections of AI-850 particles is shown in
`Fig. 3, where the bright white areas are the locations of the
`water-soluble excipients, the light gray areas are the locations
`of the paclitaxel, and the darker gray areas are the locations
`of the pores. Prior to reconstitution, the porosity of the AI-
`850 particles was predominantly internal.
`Multiple analytical methods were evaluated as options
`for quantitative analysis of the particle size distribution of the
`paclitaxel particles within a reconstituted AI-850 suspension.
`Most of these methods (e.g., laser sizing methods) require
`large dilutions to obtain particle concentrations with the op-
`erating range of the respective detector, and as a result the
`AI-850–derived paclitaxel particles dissolved before analyses
`of such dilutions could be performed. A method for quanti-
`tative analysis was developed using a Coulter Multisizer II
`
`Fig. 3. TEM of cross section of AI-850 dry powder particles.
`
`run using electrolyte saturated with paclitaxel, which prevents
`the rapid dissolution of the AI-850–derived paclitaxel par-
`ticles. The particle size distribution of the paclitaxel particles
`from a reconstituted AI-850 suspension analyzed on a Coulter
`Multisizer is shown in Fig. 4, with an observed volume mean
`particle size of 2.0 ␮m. The Coulter Multisizer method used
`an aperture that could only detect particles down to 1 ␮m.
`Thus, unlike the Malvern Mastersizer analysis of the AI-850
`microparticles, which had a lower limit of detection of 0.5 ␮m,
`the sub-micrometer particles were not detected with the
`Coulter Multisizer method, and the mean particle size value
`reported is an overestimation of the actual value. Attempts to
`perform suspension particle size analysis using a laser-based
`method on a Coulter LS230, which would have allowed for
`analysis of nano- and microparticles, using paclitaxel satu-
`rated media, were not successful due to problems attributed
`to the crystallization of paclitaxel within the paclitaxel-
`saturated media itself, affecting the LS230 analysis. An SEM
`image of paclitaxel particles isolated from a suspension post-
`reconstitution is shown in Fig. 5, which not only demonstrates
`the presence of microparticles of paclitaxel but also demon-
`strates that the microparticles themselves are internally po-
`rous, with significant surface porosity.
`Paclitaxel dissolution for the parent bulk paclitaxel lot
`and for the average of 17 lots of AI-850 is shown in Fig. 6.
`These data illustrate that the process consistently produces a
`rapidly dissolving paclitaxel product.
`
`Fig. 2. SEM of AI-850 dry powder particles.
`
`Fig. 4. Coulter Multisizer particle size distribution of reconsti-
`tuted AI-850.
`
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`351
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`MB-435. Treatment began on day 14, when the median tu-
`mor size ranged from 108 to 144 mg (individual tumors rang-
`ing from 100 mg to 172 mg). The control tumors grew well,
`with a doubling time of approximately 10 days (200–400 mg).
`Agents were administered i.v. once a day for 5 days. Maxi-
`mum weight losses observed were 4% for 15 mg/kg Taxol, and
`0%, 9%, and 21% for AI-850 at 15 mg/kg, 30 mg/kg, and 40
`mg/kg, respectively. Tumor weight data over time is shown in
`Fig. 8. At the 15 mg/kg dose, the time required for two tumor
`mass doublings was 59.6 days for Taxol and 56.9 days for
`AI-850, and the value of t-c was 44.4 days for Taxol and 41.7
`days AI-850. For 30 mg/kg dose of AI-850, the time required
`for two tumor mass doublings was 66.4 days, and the value of
`
`Fig. 5. SEM of AI-850 particles post-reconstitution.
`
`Rat Pharmacokinetic Study
`
`Plasma pharmacokinetic data for rats dosed with AI-850
`and Taxol are shown in Fig. 7 and Table I. AI-850 was well
`tolerated at all doses (5 mg/kg, 10 mg/kg, 20 mg/kg, and 30
`mg/kg of paclitaxel). Taxol administered as a 3-h infusion was
`also well tolerated at both dose levels (5 mg/kg and 10 mg/kg
`of paclitaxel). However, all the rats administered an i.v. bolus
`dose of Taxol at 10 mg/kg of paclitaxel died within 2 h.
`
`Rat Tissue Distribution Study
`
`Groups of rats were given an i.v. bolus dose of AI-850
`(5.7 mg/kg of paclitaxel) or an i.v. bolus dose of Taxol (5.7
`mg/kg of paclitaxel). Both an i.v. bolus dose of AI-850 (5.7
`mg/kg of paclitaxel) and an i.v. bolus dose of Taxol (5.7 mg/kg
`of paclitaxel) were well tolerated. Data from selected time-
`points are shown in Table II, and tissue paclitaxel AUC val-
`ues are shown in Table III.
`
`In Vivo Antitumor Efficacy
`
`To assess the efficacy of AI-850 in a tumor model, mice
`(young, adult female athymic NCr-Nu) were implanted sub-
`cutaneously (s.c.) with the human mammary tumor MDA-
`
`Fig. 6. In vitro dissolution of paclitaxel from bulk paclitaxel (䊐) and
`AI-850 (䉱). For AI-850, the error bars represent the standard devia-
`tion for 17 lots.
`
`Fig. 7. Plasma paclitaxel concentrations in Sprague-Dawley rats. All
`values are means ± standard deviations for the combined data for
`male and female rats. (a) Treatment groups were Taxol Bolus at 5
`mg/kg (䉱) and 10 mg/kg (䊏). (b) Treatment groups were Taxol In-
`fusion at 5 mg/kg (䊉) and 10 mg/kg (䊏). (c) Treatment groups
`were AI-850 Bolus at 5 mg//kg (䊊), 10 mg/kg (䊏), 20 mg/kg (䉱), and
`30 mg/kg (×).
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`Table I. Pharmacokinetic Analysis of Paclitaxel Formulations
`
`PK parameter
`
`Taxol i.v.
`bolus 5
`mg/kg
`
`Taxol i.v.
`bolus 10
`mg/kg
`
`Taxol 3-h
`infusion
`5 mg/kg
`
`Taxol 3-h
`infusion
`10 mg/kg
`
`AI-850
`i.v. bolus
`5 mg/kg
`
`AI-850
`i.v. bolus
`10 mg/kg
`
`AI-850
`i.v. bolus
`20 mg/kg
`
`AI-850
`i.v. bolus
`30 mg/kg
`
`AUCa (ng ⭈ h/ml)
`NAb
`2601 ± 386
`4427 ± 994
`NAb
`0.53 ± 0.11
`0.31 ± 0.09
`T1/2␣ (h)
`NAb
`5.98 ± 1.03
`2.80 ± 0.25
`T1/2␤ (h)
`C1 (ml h−1 kg−1)
`NAb
`2249 ± 306
`1255 ± 256
`NAb
`8363 ± 2063
`2289 ± 487
`Vdss (ml/kg)
`Cmax measured (ng/ml) 7742 ± 1763 35,338 ± 15,541 1170 ± 359
`
`12,670 ± 5741
`1.04 ± 0.25
`5.96 ± 0.83
`761 ± 231
`2085 ± 617
`6725 ± 2745
`
`6524 ± 388
`2095 ± 213
`854 ± 49
`0.77 ± 0.32
`0.40 ± 0.12
`0.24 ± 0.07
`3.28 ± 1.37
`3.18 ± 0.56
`3.49 ± 0.72
`2460 ± 154
`4388 ± 465
`5309 ± 318
`17,011 ± 2458 10,847 ± 1860 5747 ± 1094
`912 ± 116
`2140 ± 320
`4980 ± 998
`
`12,434 ± 1558
`0.62 ± 0.34
`2.50 ± 0.68
`2070 ± 259
`4778 ± 690
`8162 ± 1210
`
`a AUC for the 10 mg/kg 3-h infusion of Taxol was calculated over 0 to 24 h, whereas all the other AUC values were calculated over 0 to 12 h.
`b These data are not available for the group receiving a 10 mg/kg i.v. bolus of Taxol, as the rats died shortly after dosing.
`The values shown are the mean ± SD for combined data for male and female rats (n ⳱ 6).
`
`t-c was 51.2 days. For 40 mg/kg dose of AI-850, the time
`required for two tumor mass doublings was >76 days, and the
`value of t-c was >61 days.
`
`DISCUSSION
`AI-850 is produced via spray drying a solution containing
`paclitaxel, mannitol, polyvinylpyrrolidone C15, polysorbate
`80, and ammonium bicarbonate (the pore-forming agent).
`The pore-forming agent, ammonium bicarbonate, is removed
`during processing. Although spray drying has been described
`in the literature for a paclitaxel formulation, it has been for
`the production of a poly (lactic-co-glycolic acid)–based sus-
`tained-release microsphere formulation of paclitaxel (21),
`
`whereas AI-850 is an immediate-release formulation of pacli-
`taxel.
`The level of mannitol present in AI-850 is such that,
`post-reconstitution, AI-850 is iso-osmotic. The level of poly-
`sorbate 80 present in AI-850 (10% of the paclitaxel level)
`would result in a dose of 0.47 mg/kg polysorbate based on the
`standard dose of paclitaxel (175 mg/m2; 4.7mg/kg), which is
`well below doses of polysorbate 80 in commercial intra-
`venously administered products. For example, Taxotere
`(Aventis Pharmaceuticals, Inc., Bridgewater, NJ, USA),
`which is a similar chemotherapeutic to Taxol, is formulated with
`polysorbate 80. A standard dose of docetaxol from Taxotere
`(75 mg/m2; 2 mg/kg) would contain 52 mg/kg of polysorbate 80.
`
`Table II. Rat Tissue Distribution of Paclitaxel
`
`Organ
`
`Adrenal gland
`
`Bone marrow
`
`Brain
`
`Heart
`
`Kidney
`
`Liver
`
`Lung
`
`Muscle
`
`Plasma
`
`Sciatic nerve
`
`Small intestine
`
`Spleen
`
`Testes
`
`Thymus
`
`Agent
`
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`
`Mean (±SD) paclitaxel concentration (ng/g or ng/ml)
`
`0.5 h
`
`2 h
`
`5 h
`
`12 h
`
`13,637 ± 2068
`17,455 ± 1391
`2424a
`3034a
`112 ± 18
`161 ± 18
`8897 ± 570
`11,073 ± 2175
`14,123 ± 1086
`17,650 ± 1477
`19,102 ± 3998
`22,392 ± 5630
`14,047 ± 11,160
`11,467 ± 1396
`2867 ± 411
`3449 ± 341
`564 ± 14
`4780 ± 886
`1226 ± 333
`1860 ± 579
`11,323 ± 949
`14,698 ± 3536
`11,385 ± 1644
`13,392 ± 611
`221 ± 27
`323 ± 65
`2245 ± 733
`2552 ± 138
`
`6048 ± 1439
`6317 ± 455
`2953a
`3171a
`101 ± 32
`75 ± 12
`3767 ± 1060
`4604 ± 816
`6585 ± 1664
`7820 ± 1147
`10,630 ± 5856
`10,107 ± 2160
`9743 ± 5332
`5916 ± 1023
`2330 ± 426
`3477 ± 323
`185 ± 122
`792 ± 149
`802 ± 105
`1787 ± 347
`6270 ± 3107
`8957 ± 2562
`6960 ± 826
`8757 ± 196
`285 ± 86
`243 ± 42
`2288 ± 866
`3477 ± 323
`
`3298 ± 493
`4136 ± 1031
`2510a
`2778a
`49 ± 11
`57 ± 12
`1693 ± 226
`2499 ± 636
`3113 ± 487
`4282 ± 1697
`2887 ± 549
`7701 ± 3470
`3839 ± 3344
`2895 ± 476
`1675 ± 149
`2132 ± 154
`42 ± 5
`343 ± 170
`581 ± 61
`989 ± 306
`3003 ± 709
`5008 ± 1087
`4554 ± 811
`6174 ± 1252
`229 ± 47
`377 ± 61
`2106 ± 393
`2132 ± 154
`
`1659 ± 214
`996 ± 23
`1245a
`1440a
`36 ± 9
`BLOQ
`710 ± 40
`603 ± 10
`1414 ± 239
`847 ± 25
`1757 ± 144
`2057 ± 547
`1842 ± 390
`842 ± 86
`1024 ± 87
`905 ± 143
`18 ± 4
`43 ± 5
`345 ± 33
`311 ± 33
`1567 ± 63
`1924 ± 182
`2090 ± 86
`1578 ± 217
`181 ± 24
`237 ± 45
`2560 ± 445
`905 ± 143
`
`a Sample from all animals was pooled at each time-point due to small tissue sample size.
`
`

`
`Intravenous Hydrophobic Drug Delivery
`
`353
`
`Table III. Paclitaxel AUC in Rat Tissues
`
`Organ
`
`Adrenal gland
`
`Bone marrow
`
`Brain
`
`Heart
`
`Kidney
`
`Liver
`
`Lung
`
`Muscle
`
`Plasma
`
`Sciatic nerve
`
`Small intestine
`
`Spleen
`
`Testes
`
`Thymus
`
`Agent
`
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`AI-850
`Taxol
`
`AUC0-inf (ng⭈h/ml)
`
`66,466
`64,120
`45,817
`40,060
`1146
`1167
`34,572
`40,777
`66,516
`66,359
`80,642
`107,218
`126,782
`49,870
`38,759
`37,385
`1409
`7541
`13,016
`16,647
`67,659
`85,043
`86,871
`84,820
`19,680
`22,750
`NA
`1,432,935
`
`Two key parameters for intravenously administered sus-
`pensions of drug particles are particle size and dissolution
`rate. The particle size of reconstituted AI-850 was shown to
`be appropriate for intravenous administration, with a volume
`mean suspension particle size of less than 2.0 ␮m. The TEM
`of sections of AI-850 particles and the SEM image of the
`paclitaxel particles demonstrate the porous nature of the par-
`ticles.
`Lot to lot reproducibility of rapid in vitro dissolution of
`paclitaxel in AI-850 was observed (Fig. 6). Paclitaxel in AI-
`850 dissolved in vitro significantly more rapidly than the par-
`
`Fig. 8. Mouse MDA-MB-435 xenograft mammary tumor efficacy
`data (median tumor weight for n ⳱ 14) for the 5% dextrose control
`(⽧), 15 mg/kg Taxol (䊏), 15 mg/kg AI-850 (䉱), 30 mg/kg AI-850 (×),
`and 40 mg/kg AI-850 (䊊).
`
`ent bulk drug, with 95% of the paclitaxel in AI-850 dissolved
`in 5 min.
`Following a single i.v. bolus dose of AI-850 ranging from
`5 to 30 mg/kg of paclitaxel, plasma concentrations of pacli-
`taxel increased with increasing dose (Fig. 7c). The increases in
`Cmax were disproportionately greater than the increases in
`AI-850 doses, however, the ratio of mean measured Cmax was
`never more than 2.5 for a 2-fold increase in dose (Table I).
`AUC0-12 also increased disproportionately to dose. Both
`clearance (Cl) and steady-state volume of distribution (Vdss)
`decreased with increasing dose of AI-850, suggesting satura-
`tion of the elimination and/or distribution mechanism. Values
`for initial half-life (t1/2␣) and terminal half-life (t1/2␤) were
`consistent across the AI-850 doses tested. Overall, no differ-
`ences between the sexes were observed in the plasma phar-
`macokinetics of paclitaxel following i.v. bolus administration
`of AI-850.
`For rats receiving Taxol at 5 or 10 mg/kg of paclitaxel by
`either i.v. bolus or 3-h infusion, plasma concentrations of pac-
`litaxel increased more than proportionately with increasing
`dose (Fig. 7a,b). Doubling the i.v. bolus dose of Taxol from 5
`to 10 mg/kg of paclitaxel resulted in a 4.5-fold increase in
`mean measured Cmax at 5 min post-dose (Table I). Likewise,
`at the end of the 3-h infusion period in the 5 mg/kg and the 10
`mg/kg paclitaxel groups, plasma concentrations of paclitaxel
`were 6-fold higher for animals receiving Taxol at 10 mg/kg of
`paclitaxel than for animals receiving Taxol at 5 mg/kg of pac-
`litaxel. The Cl and Vdss of paclitaxel were lower in the 10
`mg/kg Taxol infusion group as compared to the 5 mg/kg Taxol
`infusion group. Values for terminal half-life (t1/2␤) were
`longer when Taxol was administered as a 3-h infusion than
`when administered by i.v. bolus. This difference may be due
`to either true differences in pharmacokinetics with i.v. Taxol
`bolus vs. Taxol infusion dosing or an inability to determine a
`true terminal half-life within the limitations of the sampling
`times and the assay sensitivity. Overall, no differences be-
`tween the sexes were observed in the plasma pharmacokinet-
`ics of paclitaxel following i.v. bolus or i.v. infusion adminis-
`tration of Taxol.
`A comparison of the pharmacokinetic results for AI-850
`and Taxol administered as i.v. bolus doses (Table I) demon-
`strates that at the 5 mg/kg dose, the Cl and Vdss of paclitaxel
`were 4- and 7-fold higher in those animals receiving AI-850 as
`compared to those receiving Taxol. Both t1/2␣ and t1/2␤ were
`similar across all i.v. bolus dose groups of AI-850 and Taxol.
`The paclitaxel Cmax achieved following i.v. bolus administra-
`tion of Taxol 5 mg/kg was 8-fold higher than that achieved
`with i.v. bolus administration of the same dose of AI-850. The
`data indicate that paclitaxel in AI-850 partitions more rapidly
`from the bloodstream into the tissue compartments than the
`paclitaxel in Taxol.
`Pharmacokinetic studies of Taxol in animals (28) and i