WO2020161588A1 - Process for the preparation of paclitaxel nanoparticles - Google Patents

Process for the preparation of paclitaxel nanoparticles Download PDF

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Publication number
WO2020161588A1
WO2020161588A1 PCT/IB2020/050824 IB2020050824W WO2020161588A1 WO 2020161588 A1 WO2020161588 A1 WO 2020161588A1 IB 2020050824 W IB2020050824 W IB 2020050824W WO 2020161588 A1 WO2020161588 A1 WO 2020161588A1
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Prior art keywords
paclitaxel
water
emulsion
homogenized emulsion
homogenized
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PCT/IB2020/050824
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French (fr)
Inventor
Bhaveshkumar Vallabhbhai PATEL
Rajesh Kini
Govardhan SALLA
Dhanashree MUNDHEY
Mukesh Kumar
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Orbicular Pharmaceutical Technologies Private Limited
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Publication of WO2020161588A1 publication Critical patent/WO2020161588A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/337—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having four-membered rings, e.g. taxol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein

Definitions

  • the present invention relates to a method for inhibiting the growth of paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • the invention also relates to a process for the preparation of paclitaxel
  • nanoparticles wherein the process comprising (a) preparation of paclitaxel homogenized emulsion and (b) diluting the homogenized emulsion of step (a) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
  • Paclitaxel is a natural diterpene product isolated from the pacific yew tree ( Taxus brevifolia).
  • the taxanes, paclitaxel and docetaxel belong to a novel class of anticancer drugs that stabilize microtubules and lead to tumor cell death.
  • the chemical name for paclitaxel is 5a,20-Epoxy- 1 ,2a,4,7p,10p,13a-hexahydroxytax-1 1 -en-9-one 4,10-diacetate 2-benzoate 13-ester with (2R, 3S)-N-benzoyl-3-phenylisoserine.
  • Paclitaxel has been shown to have significant antineoplastic and anticancer effects in drug-refractory ovarian cancer and has shown excellent antitumor activity in a wide variety of tumor models, and also inhibits angiogenesis when used at very low doses.
  • TAXOL® injection the commercially marketed paclitaxel injection (marketed by Bristol- Myers Squibb Co) is formulated as a concentrated nonaqueous solution in CremophorTM EL (a polyethoxylated castor oil) and dehydrated alcohol.
  • Cremophor EL improves the physical stability of the solution, and ethyl alcohol solubilizes paclitaxel.
  • the presence of Cremophor® in this formulation has been linked to severe hypersensitivity reactions in animals and humans and consequently requires premedication of individuals with corticosteroids (dexamethasone) and antihistamines.
  • ABRAXANE® The nanoparticle injectable suspension of paclitaxel was marketed as ABRAXANE® developed by Abraxis BioScience.
  • ABRAXANE® for injectable suspension paclitaxel protein-bound particles for injectable suspension
  • ABRAXANE® is supplied as a white to yellow, sterile, lyophilized powder for reconstitution with 20 ml_ of 0.9% Sodium Chloride Injection, USP prior to intravenous infusion.
  • compositions for the in vivo delivery of substantially water insoluble of substantially water insoluble
  • pharmacologically active substances such as the anticancer drug taxol
  • the pharmacologically active agent in which the pharmacologically active agent is delivered in a soluble form or in the form of suspended particles (polymeric shells) and polymeric shell having a diameter of no greater than about 10 microns.
  • It also discloses a process for the preparation of polymeric shells containing dissolved taxol wherein 5% human serum albumin solution and soybean oil used to prepare polymeric shells.
  • the process for making polymeric shells is by emulsification of the drug alone dissolved in a nonpolar solvent such as chloroform and an aqueous solution of albumin and rapidly evaporating the emulsion around 50° C.
  • U.S. Pat. No. 8,853,260 discloses a pharmaceutical formulation comprising paclitaxel at a concentration between 5 mg/ml and 15 mg/ml, wherein the pharmaceutical formulation comprises nanoparticles comprising a solid core of paclitaxel and an albumin coating with a particle size less than 400 nm.
  • the process for the preparation of paclitaxel nanoparticles comprises:
  • U.S. Pat. No. 8,728,527 discloses an injectable pharmaceutical composition comprising a substantially stable and sterile filterable dispersion of a microtubule inhibitor solid nanoparticles in an aqueous medium and its process for the preparation.
  • the process comprising combining an aqueous phase comprising water and a biocompatible polymer as emulsifier and an organic phase comprising the microtubule inhibitor, a water-immiscible organic solvent, optionally a water-miscible organic solvent as an interfacial lubricant and at least one ostwald ripening inhibitor.
  • US patent application 20050009908 discloses a process for the preparation of a stable dispersion of solid particles, in an aqueous medium comprising combining (a) a first solution comprising a substantially water- insoluble substance, a water-miscible organic solvent and an inhibitor with (b) an aqueous phase comprising water and optionally a stabilizer, thereby precipitating solid particles comprising the inhibitor and the substantially water-insoluble substance; and optionally removing the water-miscible organic solvent; wherein the inhibitor is a non-polymeric hydrophobic organic compound as defined in the description.
  • the process provides a dispersion of solid particles in an aqueous medium, which particles exhibit reduced particle growth mediated by Ostwald ripening.
  • the application describes the preparation of nanoparticles through precipitation technique using water miscible organic solvents.
  • U.S. Pat. No. 6,207,178 discloses the preparation of suspensions of colloidal solid lipid particles (SLPs) of predominantly anisometrical shape with the lipid matrix being in a stable polymorphic modification and of suspensions of micron and submicron particles of bioactive agents (PBAs).
  • SLPs colloidal solid lipid particles
  • PBAs bioactive agents
  • the present invention provides a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, by diluting the
  • the present invention provides a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • the present invention provides a process for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, wherein the process comprising (a) preparation of paclitaxel homogenized emulsion (b) diluting the homogenized emulsion of step (a) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
  • the present invention provides a process for the preparation of paclitaxel homogenized emulsion, wherein the process comprising
  • step (iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer to get homogenized emulsion of paclitaxel.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media to form a colloidal suspension of paclitaxel.
  • the colloidal suspension of paclitaxel was further subjected to solvent evaporation.
  • the colloidal suspension of paclitaxel was subjected to concentration after solvent evaporation by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators.
  • TMF tangential flow filtration
  • ultrafiltration or recirculating concentrators the colloidal suspension of paclitaxel was subjected to concentration after solvent evaporation by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators.
  • the colloidal suspension of paclitaxel was concentrated and further subjected to lyophilization.
  • homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
  • the present invention provides an injectable
  • composition comprising stable nanoparticles of paclitaxel; wherein paclitaxel nanoparticles having a mean particle size of less than about 250 nm;
  • step (iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
  • step (b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • the colloidal suspension of paclitaxel was further subjected to solvent evaporation, concentration and lyophilization.
  • homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
  • the ratio of organic phase to aqueous phase after dilution is between about 1 :60 to about 1 :150.
  • the present invention provides a method for the treatment of cancers by administering the nanosuspension composition comprising paclitaxel nanoparticles prepared in accordance with present invention.
  • Nanoparticle or Nanosuspension or colloidal suspension As used herein, the terms“Nanoparticle” or“Nanosuspension” or“colloidal
  • suspension means particles having mean particle size of less than 500 nm.
  • Nanodroplet means the emulsion droplet formed after high pressure homogenization having mean particle size of less than 500 nm.
  • the present invention relates to a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • the present invention relates to a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation before solvent evaporation.
  • the present invention also relates to a process for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, wherein the process comprising
  • step (iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
  • step (b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation before solvent evaporation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
  • the present invention provides an injectable pharmaceutical composition comprising stable nanoparticle of paclitaxel
  • step (iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
  • step (b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension.
  • step (c) colloidal suspension of step (b) was further subjected to solvent
  • step (d) colloidal suspension of step (c) was subjected to concentration by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators;
  • step (e) colloidal suspension of step (d) was further subjected to lyophilization; wherein paclitaxel nanoparticles having a mean particle size of less than about 250 nm.
  • the aqueous phase comprising water and human serum albumin may further dilute with water or aqueous media before homogenization and may optionally saturated with a sufficient amount of water immiscible solvent(s).
  • step (b) diluting the aqueous phase of step (a) with water or aqueous media and optionally saturated with a sufficient amount of water immiscible solvent(s);
  • step (d) mixing the aqueous phase of step (b) with organic phase of step (c) to form a crude emulsion
  • step (e) subjecting the crude emulsion of step (d) to homogenization using a high-pressure homogenizer to obtain homogenized emulsion;
  • step (f) homogenized emulsion of step (e) was further subjected to solvent evaporation to obtain colloidal suspension;
  • step (g) colloidal suspension of step (f) was further subjected to
  • the aqueous phase comprising water and human serum albumin.
  • Human serum albumin act as a stabilizing agent for the formation of stable nanodroplets/nanoparticles.
  • the aqueous phase comprising human serum albumin at a
  • the organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water- immiscible solvent.
  • the organic phase comprising paclitaxel at a concentration in the range of about 1 to 40% w/v.
  • the organic phase comprising paclitaxel at a concentration in the range of about 5 to 30% w/v.
  • the water immiscible solvent e.g., a solvent having less than about 5% solubility in water
  • forming an oil phase or organic phase containing paclitaxel e.g., a solvent having less than about 5% solubility in water
  • the organic phase comprising water immiscible solvent at a concentration in the range of about 1 %-99% v/v.
  • the water miscible solvent e.g., a solvent having greater than about 10% solubility in water
  • the water miscible solvent is added to the organic phase or oil phase.
  • the organic phase comprising water miscible solvent at a concentration in the range of about 1 %-99% v/v.
  • water immiscible and immiscible solvent(s), forming an oil phase or organic phase containing paclitaxel in another embodiment the water immiscible and immiscible solvent(s), forming an oil phase or organic phase containing paclitaxel.
  • water miscible and water immiscible solvents selected from, but not limited to, chloroform, methylene chloride, ethyl acetate, ethanol, tetrahydrofuran, dioxane, butanol, butyl acetate, acetonitrile, acetone, dimethyl sulfoxide, dimethyl formamide, methyl pyrrolidinone, propylene glycol, glycerol or the like combinations or mixtures thereof.
  • Additional solvents contemplated for use in the practice of the present invention include soybean oil, coconut oil, olive oil, safflower oil, cotton seed oil, sesame oil, orange oil, limonene oil, C1 -C20 alcohols, C2-C20 esters, C3- C20 ketones, polyethylene glycols, aliphatic hydrocarbons, aromatic hydrocarbons, halogenated hydrocarbons and combinations thereof.
  • the aqueous phase optionally comprising a sufficient amount of the water immiscible solvent dissolved in the aqueous phase to bring it close to the saturation concentration.
  • the aqueous and organic phase were mixed to form a crude emulsion of paclitaxel.
  • the crude emulsion of paclitaxel was further subjected to high pressure homogenization to prepare a paclitaxel homogenized emulsion.
  • Homogenized emulsion is formed by homogenization under high pressure and high shear forces. Homogenization is conveniently carried out in a high-pressure homogenizer, typically operated at pressures in the range of about 3,000 up to 60,000 psi.
  • the resulting emulsion comprises small nanodroplets of organic solvent(s) containing the dissolved paclitaxel.
  • the methods of homogenization include processes imparting high shear and cavitation such as high-pressure homogenization, high shear mixers, sonication, high shear impellers, and the like or combinations thereof.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media (dilution step) to form a colloidal suspension/ nanosuspension of paclitaxel.
  • the water immiscible solvent(s) present in the nanodroplets of homogenized emulsion Upon dilution with water or aqueous media, the water immiscible solvent(s) present in the nanodroplets of homogenized emulsion, will migrate into the water or aqueous media, which results in formation of stable colloidal suspension/ nanosuspension.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media before solvent
  • the paclitaxel homogenized emulsion produced after homogenization was diluted immediately with water or aqueous media before solvent evaporation.
  • the paclitaxel homogenized emulsion produced after homogenization was diluted with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media within 20 minutes of homogenized emulsion formation. In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media within 15 minutes of homogenized emulsion formation.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media within 10 minutes of homogenized emulsion formation.
  • the paclitaxel homogenized emulsion was diluted with water or aqueous media within 5 minutes of homogenized emulsion formation.
  • the dilution ratio of homogenized emulsion volume to water or aqueous media is between about 1 :1 to about 1 :25.
  • homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
  • the ratio of organic phase to aqueous phase after dilution is between about 1 :50 to about 1 :200.
  • the ratio of organic phase to aqueous phase after dilution is between about 1 :60 to about 1 :150.
  • the aqueous media selected from, but not limited to, normal saline solution, human serum albumin, 5% dextrose, buffer solutions and the like, or combinations thereof.
  • the diluted homogenized emulsion or colloidal suspension of paclitaxel was subjected to solvent evaporation to remove the organic solvent(s).
  • the solvent evaporation under reduced pressure yields a colloidal suspension composed of albumin bound nanoparticles of paclitaxel.
  • Acceptable methods of evaporation include the use of rotary
  • evaporators continuous flow evaporators, wiped film evaporators, flash evaporators, falling film evaporators, thin film evaporators, spray driers, freeze driers, stainless steel vessel and the like or combinations thereof.
  • the colloidal suspension of paclitaxel was subjected to concentration to remove the water or aqueous media (concentration step).
  • concentration step was performed by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators and the like.
  • the colloidal suspension was further subjected to lyophilization.
  • Colloidal suspension comprising albumin bounded nanoparticles of paclitaxel, obtained after solvent evaporation or concentration was further converted into powder form, by lyophilization or spray drying at a suitable temperature-time profile.
  • solvent evaporation, concentration and lyophilization steps were performed in a sequential order i.e. solvent evaporation followed by concentration followed by lyophilization.
  • solvent evaporation is followed by lyophilization.
  • albumin bounded nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 300 nm.
  • albumin bounded
  • nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 250 nm.
  • albumin bounded
  • nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 200 nm.
  • albumin bounded
  • nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 150 nm.
  • Nanoparticles of paclitaxel are delivered as a suspension in a biocompatible aqueous liquid.
  • This liquid may be selected from water, saline, a solution containing appropriate buffers, a solution containing nutritional agents such as amino acids, sugars, proteins, carbohydrates, vitamins or fat, and the like.
  • an injectable pharmaceutical composition comprising stable nanoparticles of paclitaxel
  • paclitaxel nanoparticles having a mean particle size of less than about 250 nm; wherein the process for preparing paclitaxel nanoparticles comprising
  • step (iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
  • step (b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation.
  • composition in one embodiment of the present invention, the composition
  • paclitaxel in an amount ranging from about 0.01 mg/ml to about 15.0 mg/ml, more preferably from about 0.1 mg/ml to about 10.0 mg/ml and most preferably from about 1.5 mg/ml to about 5.0 mg/ml.
  • paclitaxel nanoparticles prepared in accordance with present invention.
  • paclitaxel nanoparticles prepared in accordance with present invention in combination with other agents/drugs.
  • Example 1 Impact of dilution on paclitaxel particle size
  • the mixture was stirred for 5-10 min in order to obtain a crude emulsion.
  • This crude emulsion was transferred into high pressure homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained.
  • the resulting mixture was transferred to rotary evaporator and removed organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 10 min to 2 hours. This results in a colloidal suspension of paclitaxel.
  • the resulting mixture was diluted with 1000 ml_ of water under stirring. Further this resulting suspension was transferred to rotary evaporator and removed organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 10 min to 2 hours.
  • the resulting mixture was transferred to rotary evaporator and removed organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 10 min to 2 hours. This results in a colloidal suspension of paclitaxel.
  • Table 1 Impact of dilution on paclitaxel particle size
  • Example 2 Impact of dilution time point on paclitaxel particle size
  • Example 2a Impact of dilution time point on paclitaxel particle size
  • the resulting mixture was diluted with 1000 ml_ of water under stirring, immediately (To) after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
  • the paclitaxel homogenized emulsion was prepared as described above (Ex. 2a).
  • the paclitaxel homogenized emulsion was prepared as described above (Ex. 2a).
  • the paclitaxel homogenized emulsion was prepared as described above (Ex. 2a).
  • Example 3 Impact of dilution ratio of homogenized emulsion volume to water on paclitaxel particle size.
  • the paclitaxel homogenized emulsion was prepared as described above (Ex. 3a).
  • the resulting mixture was diluted with 500 ml_ of water (ratio of homogenized emulsion volume to water; 1 :5) under stirring immediately after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
  • the paclitaxel homogenized emulsion was prepared as described above (Ex. 3a).
  • the resulting mixture was diluted with 1000 ml_ of water (ratio of homogenized emulsion volume to water; 1 :10) under stirring immediately after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
  • Table 3 Impact of dilution ratio of homogenized emulsion volume to water on paclitaxel particle size.
  • Example 4 Finished product and stability details
  • Paclitaxel was dissolved in 21 .81 ml_ of chloroform and 2.09 ml_ of ethanol. The solution was added to 292.5 ml_ of 20% w/v FISA which is pre-saturated with 1 % Chloroform. The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
  • the resulting mixture was diluted with required quantity of water under stirring immediately after homogenized emulsion formation.
  • the resulting suspension was subjected to solvent evaporation in an alternate vessel where the temperature maintained between 25°C to 45°C with application of vacuum 50 mbar to 500 mbar for about 1 hr to 45 hours and further concentration using diafiltration and followed by aseptic filtration through 0.2m membrane filter, further followed by filling into vials and lyophilization.
  • Table 4 summaries the initial (To) results for the optimized formulations and Table 5 details on 6 months product stability result.

Abstract

The present invention relates to a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation. The invention also relates to a process for the preparation of paclitaxel nanoparticles, wherein the process comprising (a) preparation of paclitaxel homogenized emulsion and (b) diluting the homogenized emulsion of step (a) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.

Description

PROCESS FOR THE PREPARATION OF PACLITAXEL
NANOPARTICLES
FIELD OF THE INVENTION
The present invention relates to a method for inhibiting the growth of paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation. The invention also relates to a process for the preparation of paclitaxel
nanoparticles, wherein the process comprising (a) preparation of paclitaxel homogenized emulsion and (b) diluting the homogenized emulsion of step (a) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
BACKGROUND OF THE INVENTION
Paclitaxel is a natural diterpene product isolated from the pacific yew tree ( Taxus brevifolia). The taxanes, paclitaxel and docetaxel belong to a novel class of anticancer drugs that stabilize microtubules and lead to tumor cell death. The chemical name for paclitaxel is 5a,20-Epoxy- 1 ,2a,4,7p,10p,13a-hexahydroxytax-1 1 -en-9-one 4,10-diacetate 2-benzoate 13-ester with (2R, 3S)-N-benzoyl-3-phenylisoserine.
Paclitaxel has been shown to have significant antineoplastic and anticancer effects in drug-refractory ovarian cancer and has shown excellent antitumor activity in a wide variety of tumor models, and also inhibits angiogenesis when used at very low doses.
The poor aqueous solubility of paclitaxel, however, presents a problem for human administration. Due to this unfavorable intrinsic property, TAXOL® injection, the commercially marketed paclitaxel injection (marketed by Bristol- Myers Squibb Co) is formulated as a concentrated nonaqueous solution in Cremophor™ EL (a polyethoxylated castor oil) and dehydrated alcohol.
Cremophor EL improves the physical stability of the solution, and ethyl alcohol solubilizes paclitaxel. The presence of Cremophor® in this formulation has been linked to severe hypersensitivity reactions in animals and humans and consequently requires premedication of individuals with corticosteroids (dexamethasone) and antihistamines.
The nanoparticle injectable suspension of paclitaxel was marketed as ABRAXANE® developed by Abraxis BioScience. ABRAXANE® for injectable suspension (paclitaxel protein-bound particles for injectable suspension) is an albumin-bound form of paclitaxel with a mean particle size of approximately 130 nanometers. ABRAXANE® is supplied as a white to yellow, sterile, lyophilized powder for reconstitution with 20 ml_ of 0.9% Sodium Chloride Injection, USP prior to intravenous infusion.
U.S. Pat. Nos. 5,439,686, 5,560,933 and 5,916,596 disclose
compositions for the in vivo delivery of substantially water insoluble
pharmacologically active substances (such as the anticancer drug taxol) in which the pharmacologically active agent is delivered in a soluble form or in the form of suspended particles (polymeric shells) and polymeric shell having a diameter of no greater than about 10 microns. It also discloses a process for the preparation of polymeric shells containing dissolved taxol wherein 5% human serum albumin solution and soybean oil used to prepare polymeric shells. The process for making polymeric shells is by emulsification of the drug alone dissolved in a nonpolar solvent such as chloroform and an aqueous solution of albumin and rapidly evaporating the emulsion around 50° C.
U.S. Pat. No. 8,853,260 discloses a pharmaceutical formulation comprising paclitaxel at a concentration between 5 mg/ml and 15 mg/ml, wherein the pharmaceutical formulation comprises nanoparticles comprising a solid core of paclitaxel and an albumin coating with a particle size less than 400 nm. The process for the preparation of paclitaxel nanoparticles
comprising combining 3% HSA solution with paclitaxel solution (prepared by using chloroform/ ethyl alcohol mixture) and mix the paclitaxel/HSA solution. The mixed solution was poured into the chamber of the homogenizer and cycle the mixture through the homogenizer at a pressure until the desired particle size of polymeric shells was obtained. The milky suspension of polymeric shells containing dissolved paclitaxel were diluted to 10 ml in normal saline. This suspension was placed in a rotary evaporator and the volatile organic removed by vacuum.
U.S. Pat. No. 8,728,527 discloses an injectable pharmaceutical composition comprising a substantially stable and sterile filterable dispersion of a microtubule inhibitor solid nanoparticles in an aqueous medium and its process for the preparation. The process comprising combining an aqueous phase comprising water and a biocompatible polymer as emulsifier and an organic phase comprising the microtubule inhibitor, a water-immiscible organic solvent, optionally a water-miscible organic solvent as an interfacial lubricant and at least one ostwald ripening inhibitor.
US patent application 20050009908 discloses a process for the preparation of a stable dispersion of solid particles, in an aqueous medium comprising combining (a) a first solution comprising a substantially water- insoluble substance, a water-miscible organic solvent and an inhibitor with (b) an aqueous phase comprising water and optionally a stabilizer, thereby precipitating solid particles comprising the inhibitor and the substantially water-insoluble substance; and optionally removing the water-miscible organic solvent; wherein the inhibitor is a non-polymeric hydrophobic organic compound as defined in the description. The process provides a dispersion of solid particles in an aqueous medium, which particles exhibit reduced particle growth mediated by Ostwald ripening. The application describes the preparation of nanoparticles through precipitation technique using water miscible organic solvents.
U.S. Pat. No. 6,207,178 discloses the preparation of suspensions of colloidal solid lipid particles (SLPs) of predominantly anisometrical shape with the lipid matrix being in a stable polymorphic modification and of suspensions of micron and submicron particles of bioactive agents (PBAs). A suspension stable for at least about 12 months of particles of bioactive agents (PBAs) manufactured by an emulsifying process having the following steps: (a) melting at least one solid bioactive agent; (b) heating a dispersion medium to approximately the same temperature as said at least one molten solid bioactive agent formed by step (a); (c) adding at least one highly mobile water-soluble or dispersible stabilizer, which does not form a separate phase in the dispersion medium, to the dispersion medium in an amount effective after emulsification to stabilize newly created surfaces during recrystallization, and optionally adding at least one lipid-soluble or dispersible stabilizer to said at least one molten bioactive agent; (d) premixing said at least one molten bioactive agent and the dispersion medium, and subsequently homogenizing said mixture by high pressure homogenization, micro-fluidization and/or ultrasonication; and (e) allowing the homogenized dispersion to cool until solid particles are formed by recrystallization of the dispersed bioactive agents.
An increase in the paclitaxel droplet size after homogenization was observed, which results in, increase in the paclitaxel nanoparticles size after solvent evaporation with the methods disclosed in the prior arts.
Hence, it is the objective of the present invention to develop a method or process to prevent the growth of paclitaxel droplet size in the homogenized emulsions, during the preparation of paclitaxel nanoparticles.
SUMMARY OF THE INVENTION
The present invention provides a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, by diluting the
homogenized emulsion with water or aqueous media.
In one aspect, the present invention provides a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation.
In another aspect, the present invention provides a process for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, wherein the process comprising (a) preparation of paclitaxel homogenized emulsion (b) diluting the homogenized emulsion of step (a) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
In another aspect, the present invention provides a process for the preparation of paclitaxel homogenized emulsion, wherein the process comprising
(i) mixing an aqueous phase comprising water and human serum albumin; with (ii) an organic phase comprising paclitaxel, at least one water- miscible solvent and at least one water-immiscible solvent to form a crude emulsion; and
(iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer to get homogenized emulsion of paclitaxel.
In another aspect of present invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media to form a colloidal suspension of paclitaxel.
In another aspect of present invention, the colloidal suspension of paclitaxel was further subjected to solvent evaporation.
In another aspect of present invention, the colloidal suspension of paclitaxel was subjected to concentration after solvent evaporation by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators.
In another aspect of present invention, the colloidal suspension of paclitaxel was concentrated and further subjected to lyophilization.
In another aspect of present invention, the dilution ratio of
homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
In one aspect, the present invention provides an injectable
pharmaceutical composition comprising stable nanoparticles of paclitaxel; wherein paclitaxel nanoparticles having a mean particle size of less than about 250 nm;
wherein the process for preparing paclitaxel nanoparticles comprising
(a) preparation of paclitaxel homogenized emulsion by
(i) mixing an aqueous phase comprising water and human serum albumin; with
(ii) an organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water-immiscible solvent to form a crude emulsion; and
(iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
(b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation. In another aspect of present invention, the colloidal suspension of paclitaxel was further subjected to solvent evaporation, concentration and lyophilization.
In another aspect of present invention, the dilution ratio of
homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
In another aspect of present invention, the ratio of organic phase to aqueous phase after dilution is between about 1 :60 to about 1 :150.
In one aspect, the present invention provides a method for the treatment of cancers by administering the nanosuspension composition comprising paclitaxel nanoparticles prepared in accordance with present invention.
DETAILED DESCRIPTION OF THE INVENTION
Nanoparticle or Nanosuspension or colloidal suspension. As used herein, the terms“Nanoparticle” or“Nanosuspension” or“colloidal
suspension” means particles having mean particle size of less than 500 nm.
Nanodroplet. As used herein, the term“Nanodroplet” means the emulsion droplet formed after high pressure homogenization having mean particle size of less than 500 nm.
The present invention relates to a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation.
The present invention relates to a method for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, during the preparation of paclitaxel nanoparticle, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation before solvent evaporation.
The present invention also relates to a process for inhibiting the growth of the paclitaxel droplet size in a homogenized emulsion, wherein the process comprising
(a) preparation of paclitaxel homogenized emulsion by (i) mixing an aqueous phase comprising water and human serum albumin; with
(ii) an organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water-immiscible solvent to form a crude emulsion; and
(iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
(b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation before solvent evaporation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
The present invention provides an injectable pharmaceutical composition comprising stable nanoparticle of paclitaxel;
wherein the process for preparing paclitaxel nanoparticle comprising
(a) preparation of paclitaxel homogenized emulsion by
(i) mixing an aqueous phase comprising water and human serum albumin; with
(ii) an organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water-immiscible solvent to form a crude emulsion; and
(iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
(b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension.
(c) colloidal suspension of step (b) was further subjected to solvent
evaporation;
(d) colloidal suspension of step (c) was subjected to concentration by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators;
(e) colloidal suspension of step (d) was further subjected to lyophilization; wherein paclitaxel nanoparticles having a mean particle size of less than about 250 nm.
In one embodiment of the invention, the aqueous phase comprising water and human serum albumin may further dilute with water or aqueous media before homogenization and may optionally saturated with a sufficient amount of water immiscible solvent(s).
In another embodiment of the invention, the process for preparing paclitaxel nanoparticle comprising
(a) preparation of an aqueous phase comprising water and human serum albumin;
(b) diluting the aqueous phase of step (a) with water or aqueous media and optionally saturated with a sufficient amount of water immiscible solvent(s);
(c) preparation of an organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water-immiscible solvent to form a crude emulsion
(d) mixing the aqueous phase of step (b) with organic phase of step (c) to form a crude emulsion;
(e) subjecting the crude emulsion of step (d) to homogenization using a high-pressure homogenizer to obtain homogenized emulsion;
(f) homogenized emulsion of step (e) was further subjected to solvent evaporation to obtain colloidal suspension;
(g) colloidal suspension of step (f) was further subjected to
lyophilization.
In one embodiment of the invention the aqueous phase comprising water and human serum albumin. Human serum albumin act as a stabilizing agent for the formation of stable nanodroplets/nanoparticles.
The aqueous phase comprising human serum albumin at a
concentration in the range of about 0.05 to 25% (w/v).
In one embodiment of the invention, the organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water- immiscible solvent.
In one embodiment, the organic phase comprising paclitaxel at a concentration in the range of about 1 to 40% w/v.
In one embodiment, the organic phase comprising paclitaxel at a concentration in the range of about 5 to 30% w/v. The water immiscible solvent (e.g., a solvent having less than about 5% solubility in water) at high concentration, forming an oil phase or organic phase containing paclitaxel.
In one embodiment of the invention, the organic phase comprising water immiscible solvent at a concentration in the range of about 1 %-99% v/v.
The water miscible solvent (e.g., a solvent having greater than about 10% solubility in water) is added to the organic phase or oil phase.
In one embodiment of the invention, the organic phase comprising water miscible solvent at a concentration in the range of about 1 %-99% v/v.
In another embodiment the water immiscible and immiscible solvent(s), forming an oil phase or organic phase containing paclitaxel.
In one embodiment of the invention, water miscible and water immiscible solvents selected from, but not limited to, chloroform, methylene chloride, ethyl acetate, ethanol, tetrahydrofuran, dioxane, butanol, butyl acetate, acetonitrile, acetone, dimethyl sulfoxide, dimethyl formamide, methyl pyrrolidinone, propylene glycol, glycerol or the like combinations or mixtures thereof.
Additional solvents contemplated for use in the practice of the present invention include soybean oil, coconut oil, olive oil, safflower oil, cotton seed oil, sesame oil, orange oil, limonene oil, C1 -C20 alcohols, C2-C20 esters, C3- C20 ketones, polyethylene glycols, aliphatic hydrocarbons, aromatic hydrocarbons, halogenated hydrocarbons and combinations thereof.
In one embodiment of the invention, the aqueous phase optionally comprising a sufficient amount of the water immiscible solvent dissolved in the aqueous phase to bring it close to the saturation concentration.
In one embodiment of the invention, the aqueous and organic phase were mixed to form a crude emulsion of paclitaxel.
In another embodiment of the invention, the crude emulsion of paclitaxel was further subjected to high pressure homogenization to prepare a paclitaxel homogenized emulsion.
Homogenized emulsion is formed by homogenization under high pressure and high shear forces. Homogenization is conveniently carried out in a high-pressure homogenizer, typically operated at pressures in the range of about 3,000 up to 60,000 psi. The resulting emulsion comprises small nanodroplets of organic solvent(s) containing the dissolved paclitaxel.
In one embodiment of the invention, the methods of homogenization include processes imparting high shear and cavitation such as high-pressure homogenization, high shear mixers, sonication, high shear impellers, and the like or combinations thereof.
In one embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media (dilution step) to form a colloidal suspension/ nanosuspension of paclitaxel.
Upon dilution with water or aqueous media, the water immiscible solvent(s) present in the nanodroplets of homogenized emulsion, will migrate into the water or aqueous media, which results in formation of stable colloidal suspension/ nanosuspension.
The migration of water immiscible solvent(s) from the nanodroplets of homogenized emulsion to water or aqueous media will prevent the growth of nanodroplet size which in turn control or stabilize the paclitaxel nanoparticles size before solvent evaporation.
In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media before solvent
evaporation.
In one embodiment of the invention, the paclitaxel homogenized emulsion produced after homogenization was diluted immediately with water or aqueous media before solvent evaporation.
In one embodiment of the invention, the paclitaxel homogenized emulsion produced after homogenization was diluted with water or aqueous media within 30 minutes of homogenized emulsion formation.
In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media within 30 minutes of homogenized emulsion formation.
In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media within 20 minutes of homogenized emulsion formation. In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media within 15 minutes of homogenized emulsion formation.
In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media within 10 minutes of homogenized emulsion formation.
In another embodiment of the invention, the paclitaxel homogenized emulsion was diluted with water or aqueous media within 5 minutes of homogenized emulsion formation.
In one embodiment of the invention, the dilution ratio of homogenized emulsion volume to water or aqueous media is between about 1 :1 to about 1 :25.
In another embodiment of the invention, the dilution ratio of
homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
In one embodiment of the invention, the ratio of organic phase to aqueous phase after dilution is between about 1 :50 to about 1 :200.
In another embodiment of the invention, the ratio of organic phase to aqueous phase after dilution is between about 1 :60 to about 1 :150.
In another embodiment of the invention, the aqueous media selected from, but not limited to, normal saline solution, human serum albumin, 5% dextrose, buffer solutions and the like, or combinations thereof.
In one embodiment of the invention, the diluted homogenized emulsion or colloidal suspension of paclitaxel was subjected to solvent evaporation to remove the organic solvent(s).
The solvent evaporation under reduced pressure yields a colloidal suspension composed of albumin bound nanoparticles of paclitaxel.
Acceptable methods of evaporation include the use of rotary
evaporators, continuous flow evaporators, wiped film evaporators, flash evaporators, falling film evaporators, thin film evaporators, spray driers, freeze driers, stainless steel vessel and the like or combinations thereof.
In another embodiment of the invention, the colloidal suspension of paclitaxel was subjected to concentration to remove the water or aqueous media (concentration step). The above step was performed by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators and the like.
In one embodiment of the invention, after concentration step, the colloidal suspension was further subjected to lyophilization.
Colloidal suspension comprising albumin bounded nanoparticles of paclitaxel, obtained after solvent evaporation or concentration was further converted into powder form, by lyophilization or spray drying at a suitable temperature-time profile.
In one embodiment of the invention, solvent evaporation, concentration and lyophilization steps were performed in a sequential order i.e. solvent evaporation followed by concentration followed by lyophilization.
In another embodiment of the invention, solvent evaporation is followed by lyophilization.
In one embodiment of the invention, albumin bounded nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 300 nm.
In another embodiment of the invention, albumin bounded
nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 250 nm.
In another embodiment of the invention, albumin bounded
nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 200 nm.
In another embodiment of the invention, albumin bounded
nanoparticles of paclitaxel prepared in accordance with present invention having a mean particle size of less than about 150 nm.
Nanoparticles of paclitaxel are delivered as a suspension in a biocompatible aqueous liquid. This liquid may be selected from water, saline, a solution containing appropriate buffers, a solution containing nutritional agents such as amino acids, sugars, proteins, carbohydrates, vitamins or fat, and the like.
In one embodiment of the invention, an injectable pharmaceutical composition comprising stable nanoparticles of paclitaxel;
wherein paclitaxel nanoparticles having a mean particle size of less than about 250 nm; wherein the process for preparing paclitaxel nanoparticles comprising
(a) preparation of paclitaxel homogenized emulsion by
(i) mixing an aqueous phase comprising water and human serum albumin; with
(ii) an organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water-immiscible solvent to form a crude emulsion; and
(iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
(b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation.
In one embodiment of the present invention, the composition
comprising paclitaxel in an amount ranging from about 0.01 mg/ml to about 15.0 mg/ml, more preferably from about 0.1 mg/ml to about 10.0 mg/ml and most preferably from about 1.5 mg/ml to about 5.0 mg/ml.
In one embodiment of the invention, provide a method for the treatment of cancers by administering the nanosuspension composition comprising paclitaxel nanoparticles prepared in accordance with present invention.
In another embodiment of the invention, provide a method for the treatment of cancers by administering the nanosuspension composition comprising paclitaxel nanoparticles prepared in accordance with present invention in combination with other agents/drugs.
To further illustrate the invention, the following examples are provided.
It is to be understood that these examples are provided for illustrative purposes and are not to be construed as limiting the scope of the present invention in any manner whatsoever.
Example 1 : Impact of dilution on paclitaxel particle size
Example 1a:
2 g of Paclitaxel was dissolved in 6.75 ml_ of chloroform and 0.65 ml_ of Ethanol. The solution was added to 90 ml_ of 20% w/v HSA which is pre saturated with 1 % Chloroform.
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained.
The resulting mixture was transferred to rotary evaporator and removed organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 10 min to 2 hours. This results in a colloidal suspension of paclitaxel.
Example 1 b:
2 g of Paclitaxel was dissolved in 6.75 ml_ of chloroform and 0.65 ml_ of ethanol. The solution was added to 90 ml_ of 20% w/v HSA which is pre saturated with 1 % chloroform.
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained.
The resulting mixture was diluted with 1000 ml_ of water under stirring. Further this resulting suspension was transferred to rotary evaporator and removed organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 10 min to 2 hours.
Example 1c:
90 ml_ of 20% w/v HSA solution was taken and diluted to 1 100 ml_ with water. 2 g of Paclitaxel was dissolved in 6.75 ml_ of chloroform and 0.65 ml_ of ethanol, this solution was added to above diluted albumin solution.
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained.
The resulting mixture was transferred to rotary evaporator and removed organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 10 min to 2 hours. This results in a colloidal suspension of paclitaxel.
Table 1 : Impact of dilution on paclitaxel particle size
Figure imgf000015_0001
Example 2: Impact of dilution time point on paclitaxel particle size Example 2a:
2 g of Paclitaxel was dissolved in 6.75 imL of chloroform and 0.65 ml_ of ethanol. The solution was added to 90 ml_ of 20% w/v HSA which is pre saturated with 1 % chloroform.
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained.
The resulting mixture was diluted with 1000 ml_ of water under stirring, immediately (To) after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Example 2b:
As another example, the paclitaxel homogenized emulsion was prepared as described above (Ex. 2a).
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained. The resultant mixture was collected and kept under stirring at 2-8° C.
After 5 minutes of collection, the mixture was diluted with 1000 ml_ of water under stirring. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Example 2c:
As another example, the paclitaxel homogenized emulsion was prepared as described above (Ex. 2a).
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained. The resultant mixture was collected and kept under stirring at 2-8° C. After 15 minutes of collection, the mixture was diluted with 1000 ml_ of water under stirring. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Example 2d:
As another example, the paclitaxel homogenized emulsion was prepared as described above (Ex. 2a).
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired droplet size was obtained. The resultant mixture was collected and kept under stirring at 2-8° C.
After 30 minutes of collection, the mixture was diluted with 1000 ml_ of water under stirring. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Table 2: Impact of dilution time point on paclitaxel particle size
Figure imgf000017_0001
Example 3: Impact of dilution ratio of homogenized emulsion volume to water on paclitaxel particle size.
Example 3a:
2 g of Paclitaxel was dissolved in 6.75 ml_ of chloroform and 0.65 ml_ of ethanol. The solution was added to 90 ml_ of 20% w/v FISA which is pre saturated with 1 % chloroform.
The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until desired size was obtained. The resulting mixture was diluted with 200 ml_ of water (ratio of homogenized emulsion volume to water; 1 :2) under stirring immediately after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Example 3b:
As another example, the paclitaxel homogenized emulsion was prepared as described above (Ex. 3a).
The resulting mixture was diluted with 500 ml_ of water (ratio of homogenized emulsion volume to water; 1 :5) under stirring immediately after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Example 3c:
As another example, the paclitaxel homogenized emulsion was prepared as described above (Ex. 3a).
The resulting mixture was diluted with 1000 ml_ of water (ratio of homogenized emulsion volume to water; 1 :10) under stirring immediately after homogenized emulsion formation. Further this resulting suspension was transferred to rotary evaporator and removed the organic solvents at 40-50°C, at reduced pressure 100-200 mbar for 1 to 2 hours.
Table 3: Impact of dilution ratio of homogenized emulsion volume to water on paclitaxel particle size.
Figure imgf000018_0001
Example 4: Finished product and stability details
6.5 g of Paclitaxel was dissolved in 21 .81 ml_ of chloroform and 2.09 ml_ of ethanol. The solution was added to 292.5 ml_ of 20% w/v FISA which is pre-saturated with 1 % Chloroform. The mixture was stirred for 5-10 min in order to obtain a crude emulsion. This crude emulsion was transferred into high pressure
homogenizer and processed at pressure 1000 to 35000 psi until the desired particle size of <250 nm obtained.
The resulting mixture was diluted with required quantity of water under stirring immediately after homogenized emulsion formation. The resulting suspension was subjected to solvent evaporation in an alternate vessel where the temperature maintained between 25°C to 45°C with application of vacuum 50 mbar to 500 mbar for about 1 hr to 45 hours and further concentration using diafiltration and followed by aseptic filtration through 0.2m membrane filter, further followed by filling into vials and lyophilization. Table 4 summaries the initial (To) results for the optimized formulations and Table 5 details on 6 months product stability result.
Table 4: Stability details of initial (To)
Figure imgf000019_0001
Table 5: Product stability data of 6 months
Figure imgf000019_0002
Figure imgf000020_0001

Claims

We Claim,
1. A method for inhibiting the growth of the paclitaxel emulsion droplet size in the homogenized emulsion, by diluting the homogenized emulsion with water or aqueous media within 30 minutes of homogenized emulsion formation.
2. A process for inhibiting the growth of the paclitaxel droplet size in the homogenized emulsion, wherein the process comprising
(a) preparation of paclitaxel homogenized emulsion by
(i) mixing an aqueous phase comprising water and human serum albumin; with
(ii) an organic phase comprising paclitaxel, at least one water-miscible solvent and at least one water-immiscible solvent to form a crude emulsion; and
(iii) subjecting the crude emulsion of step (ii) to homogenization using a high-pressure homogenizer;
(b) diluting the homogenized emulsion of step (a) (iii) with water or aqueous media within 30 minutes of homogenized emulsion formation to get a colloidal suspension of paclitaxel having a mean particle size less than about 250 nm.
3. The process according to the claim 2, the colloidal suspension of step (b) was subjected to solvent evaporation.
4. The process according to the claim 3, the colloidal suspension was subjected to concentration, by using tangential flow filtration (TFF) or ultrafiltration or recirculating concentrators.
5. The process according to the claim 4, the colloidal suspension was further subjected to lyophilization.
6. The process according to the claim 2, wherein the dilution ratio of homogenized emulsion volume to water or aqueous media is between about 1 :5 to about 1 :20.
7. The process according to claim 2, wherein water-miscible and water immiscible solvents selected from, but not limited to, chloroform, methylene chloride, ethyl acetate, ethanol, tetrahydrofuran, dioxane, butanol, butyl acetate, acetonitrile, acetone, dimethyl sulfoxide, dimethyl formamide, methyl pyrrolidinone, combinations or mixtures thereof.
8. The process according to claim 1 , wherein aqueous media selected from, but not limited to, normal saline solution, human serum albumin, buffer solutions.
9. The process according to claim 2, wherein human serum albumin is present at a concentration in the range of about 0.05 to 25% (w/v).
10. The process according to claim 3, wherein the methods of evaporation include the use of rotary evaporators, continuous flow
evaporators, wiped film evaporators, flash evaporators, falling film
evaporators, thin film evaporators, spray driers, freeze driers, tangential flow filtration, and the like.
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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2015018380A2 (en) * 2014-07-03 2015-02-12 Cspc Zhongqi Pharmaceutical Technology(Shijiazhuang)Co., Ltd. Therapeutic nanoparticles and the preparation methods thereof
US20150104521A1 (en) * 1997-06-27 2015-04-16 Abraxis Bioscience, Llc Novel formulations of pharmacological agents, methods for the preparation thereof and methods for the use thereof
US20150111960A1 (en) * 1996-10-01 2015-04-23 Abraxis Bioscience, Llc Novel formulations of pharmacological agents, methods for the preparation thereof and methods for the use thereof

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20150111960A1 (en) * 1996-10-01 2015-04-23 Abraxis Bioscience, Llc Novel formulations of pharmacological agents, methods for the preparation thereof and methods for the use thereof
US20150104521A1 (en) * 1997-06-27 2015-04-16 Abraxis Bioscience, Llc Novel formulations of pharmacological agents, methods for the preparation thereof and methods for the use thereof
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