DESC:FIELD OF THE INVENTION
The present invention relates to stable paclitaxel and albumin nanosuspension suitable for intravenous administration and a method of preparation thereof.
BACKGROUND OF THE INVENTION
Cancer is one of the leading causes of deaths worldwide with an estimated 7.6 million individuals lost each year and accounting for 13% of all deaths. Cancer is a single disease which affects a multitude of organs with each organ or system developing a distinct set of symptoms and methods for treating it. Many instances of cancer could be avoided, with some estimates indicating that about 30% of cancer deaths are associated with smoking or other lifestyle factors or dietary practices that could potentially be avoided by changes in human behavior. Nonetheless, majority of cancers cannot be avoided by simple behavioral changes and require technological innovation to improve outcomes. The success in limiting cancer could be further enhanced by more widespread development and implementation of these advanced technologies.
Nanotechnology and nanoscience methods are increasingly being evaluated as a potential solution for treatment of cancer. Nanotechnology is a multidisciplinary area of inquiry and application. Nanosuspensions are colloidal dispersions of nanosized drug particles stabilized by surfactants. They can also be defined as a biphasic system or carrier-free colloidal drug delivery system consisting of pure drug particles and stabilizers dispersed in an aqueous or non-aqueous vehicle in which the diameter of the suspended particle is less than 1µm in size. The reduced particle size in the nanosuspension technology obviates the possibility of blood capillaries blockade by intravenous administration rendering it a safe method for administration of poorly soluble drugs.
Paclitaxel, a poorly soluble drug, was first isolated in 1971 from the Pacific yew and approved for medical use in 1993. It is listed on the World Health Organization's list of Essential Medicines, as the most effective and safe medicine needed in a health system. Worldwide, it is approved for treatment of ovarian, breast, lung, bladder, prostate, melanoma, esophageal, and other types of solid tumor cancers as well as Kaposi's sarcoma.
Due to its aqueous insolubility, paclitaxel was first formulated by using a mixture of Cremophor (50%) and absolute alcohol (50%) for the use of parenteral administration. Cremophor EL, as a formulation vehicle, is not inert biologically and pharmacologically. It is involved with toxic side effects such as hypersensitivity reaction, anaphylactic shock, neuropathy, ‘pseudo’- nonlinear pharmacokinetic behavior of paclitaxel and death. With this formulation, premedication with steroids, antihistamines and/or H2 receptor blockers was required prior to dosing in order to prevent hypersensitivity reactions. Additionally, specialized IV tubings were required to prevent leaching of plasticizers.
Albumin bound paclitaxel, is an improved alternative formulation where paclitaxel is bound to albumin in the form of nanosuspension and is marketed worldwide under the brand name of ABRAXANE®. U.S. Patent No. 7,820,788 discloses that the pharmaceutical composition is in form of nanosuspension for intravenous administration comprising paclitaxel and albumin and the particle size of the formulation is not more than 200 nm. Albumin bound nanoparticles are biocompatible, non-immunogenic and biodegradable.
Albumin is useful for targeting cancer cells as it is taken up by growing tumor cells as a source of amino acids and energy due to increasing cellular stress. Moreover, the specificity of human serum albumin (HSA) for glycoprotein 60 (gp60) receptor present on the surface of tumor cells allows target specific delivery, thus reducing immune response. Albumin bound nature of the formulation also improves therapeutic outcome by enhanced permeability and retention (EPR) effect as compared to free drug administration. Additionally, the formulation is solvent free and hence does not cause hypersensitivity reaction and permits administration of higher dose without any premedication.
In the marketed ABRAXANE® formulation, the weight ratio of albumin to paclitaxel is about 9:1. Further, for ABRAXANE® formulation the available literature suggests that the albumin/paclitaxel ratio of about 9:1 is associated with higher therapeutic efficacy and substantially reduced toxicity when compared with the formulation that differs in the ratio of albumin to paclitaxel. The formulation having albumin to paclitaxel ratio 9:1 has increased cellular binding, therefore slight variation from this ratio is likely to affect the safety and efficacy profile of the formulation.
It is needless to mention that the slight variation in the manufacturing process may change the ratio of albumin to paclitaxel available in the final product. In fact, the inventors of the ABRAXANE® formulation have reported that during the manufacturing of the formulation, the starting ratio of albumin to drug always increased in the final resultant composition, i.e. the final ratio was 13.3:1, when the starting ratio was 9:1. Not only with the manufacturing process, but inventors have also reported that even during scaling up, the ratio increases.
The principal parameters of nanoparticle formulation are their shape, size, and the morphological sub-structure of the substance. Their small size and large surface area can lead to particle-particle agglomeration, making physical handling of nanoparticles difficult in liquid and dry forms. In addition, small particles size and large surface area result in limited drug loading and burst release.
Existing manufacturing processes from the literature describe solvent removal by rotary evaporation immediately after particle size reduction step. However, in this process, the solvent load on the evaporation apparatus is very high. Therefore, this leads to lengthy process and high levels of residual solvents in the finished product. The extent of residual solvent present in the system can adversely affect particle agglomeration, may lead to particle size variation and high polydispersity index (PDI).
Therefore, there is a need of a manufacturing process, which is not dependent on critical vagaries of the process, such as variation in paclitaxel to albumin ratio, the scale of the preparation, shape and size of the nanoparticles and particle agglomeration.
Thus, the objective of the present invention is to maintain the desired particle size of paclitaxel-albumin nanoparticle, during the course of manufacturing process. Another objective is to increase physical and chemical stability, by reducing the level of physiologically harmful organic solvents. It is desirable that such pharmaceutical compositions should remain stable under conventional storage conditions and remain physically and/or chemically stable after rehydration or reconstitution.

SUMMARY OF THE INVENTION
The present invention is directed towards a stable paclitaxel and albumin nanosuspension for intravenous administration.
The present invention is directed towards a process for preparation of a stable paclitaxel and albumin nanosuspension comprising:
a) preparing aqueous phase by dissolving human albumin in water
b) preparing non-aqueous phase by dissolving paclitaxel in organic solvent
c) preparation of crude emulsion of (a) & (b)
d) preparation of nanosuspension of (c)
e) dilution of nanosuspension of (d) with water
f) removal of solvents
g) aseptic filtration
h) filling, followed by
i) lyophilization
The process of the present invention has no impact on the particle size of paclitaxel- albumin nanoparticle.
DETAILED DESCRIPTION OF THE INVENTION
The present invention is directed towards a stable paclitaxel and albumin nanosuspension for intravenous administration.
In one of the embodiment, the process for preparation of a stable paclitaxel and albumin nanosuspension according to the present invention typically involves the following steps:
a) preparing aqueous phase by dissolving human albumin in water
b) preparing non-aqueous phase by dissolving paclitaxel in organic solvent
c) preparation of crude emulsion of (a) & (b)
d) preparation of nanosuspension of (c)
e) dilution of nanosuspension of (d) with water
f) removal of solvent
g) aseptic filtration
h) filling followed by
i) lyophilization
In conventional methods for preparation of paclitaxel and albumin nanosuspension, the formed nanoparticle is exposed to organic solvents for a prolonged period. Hence albumin desolvation occurs causing paclitaxel-albumin nanoparticles to coalesce and form large agglomerate, subsequently increasing the particle size and PDI. Further, in the conventional rotary evaporation process, achieving set temperature and vacuum level, which is critical, requires considerable amount of time which lengthens the process, thereby making it economically, non-viable.
The present inventors have found that the immediate dilution of bulk solution with water, after size reduction step (d) helps in reducing the effective concentration of organic solvent and, in turn, avoids destabilization of paclitaxel- albumin nanoparticle.
The term "nanoparticle" or "nanoparticulate" as used herein refers to the solid particles of active ingredient having a particle size in nanometer (nm), such that 90% of the particles (D90) have a size not more than 1000nm. It may be noted that the solid particles may include excipients adsorbed onto its surface, such as for example wetting agents, surfactants or surface stabilizers, which are adsorbed onto the surface of the active ingredient and there is no composite particle formed thereof.
The term "nanosuspension" refers to a suspension obtained after size reduction, comprising the solid particles of paclitaxel and albumin having particle size in nm, such that 90% of the particles (D90) have a size not more than 1000nm.
Typically, the pharmaceutically stable composition of the present invention remains physically and/or chemically stable without any precipitation or sedimentation for at least about 8 hours, or up to about 24 hours following reconstitution or rehydration when stored in refrigerator (2 to 8ºC) and/or at room temperature (20 to 25ºC). The stable nanosuspension of the present invention obviates the formation of floccules and maintains the particle size in an acceptable range of not more than 1000nm, preferably not more than 500nm, more preferably not more than 230nm (Table 3).
In the present invention the organic solvent is used to suspend or dissolve paclitaxel. The organic solvent may include aliphatic, cycloaliphatic, or aromatic hydrocarbons having 4-30 carbon atoms such as n-dodecane, n-decane, n-hexane, cyclohexane, toluene, benzene, and mixtures thereof, aliphatic or aromatic alcohols having 2-30 carbon atoms such as ethanol, octanol and mixtures thereof, alkyl, aryl, or cyclic ethers having 2-30 carbon atoms such as diethyl ether, tetrahydrofuran, and mixtures thereof, alkyl or aryl halides having 1-30 carbon atoms selected from the group of CHCl3, CH2Cl2,CH3ClCH2Cl2, CH2Cl-CH2Cl, and mixtures thereof, aliphatic or aromatic esters having 2-30 carbon atoms such as ethyl acetate, methyl acetate, ethyl caprylate (octanoate) and mixtures thereof , ketones having 3-30 carbon atoms such as acetone, methyl ethyl ketone and mixtures thereof, polyalkylene glycols such as polyethylene glycol, and mixtures thereof.
The preferred organic solvent(s) is chloroform, ethanol, or mixtures thereof.
The crude emulsion is prepared using homogenization or mixing or sonication method while the nanosuspension is prepared by high pressure homogenizer or ultrasonication or extrusion or high energy wet milling techniques.
During manufacturing process, the nanosuspension obtained after step (d) is diluted with water in an amount that is 2 to 10 times the quantity of nanosuspension bulk of step (d), preferably 2 to 8 times the quantity of nanosuspension bulk of step (d), preferably 2 to 6 times the quantity of nanosuspension bulk of step (d), preferably 2 to 4 times the quantity of nanosuspension bulk of step (d).
The diluted nanosuspension is passed through tangential flow filtration or wiped film evaporator or thin film evaporator or combinations thereof for removal of organic solvents and excess water. After final concentration, small amounts of nanosuspension are transferred to rotary evaporator for removal of residual organic solvents under reduced pressure.
Accordingly, limiting the exposure to organic solvents prevents the paclitaxel-albumin nanoparticle from agglomerate formation, thus maintains the particles in acceptable size range of not more than 230nm. Hence immediate removal of organic solvent/s results in desired particle size.
The following examples further illustrate the invention, but should not be construed as in any way limiting its scope. In particular, the processing conditions are merely exemplary and can be readily varied by one of ordinary skill in the art.
EXAMPLES
Paclitaxel and albumin nanosuspension were made by using composition as summarized in below Table 1.
Table 1: Composition of paclitaxel and albumin nanosuspension
Sr. No. Ingredients Formulation 1 Formulation 2 Formulation
3
1 Paclitaxel 100 mg
2 Human albumin Approximately 900mg
4 Chloroform q.s. q.s. --
5 Ethanol -- q.s. q.s.
6 Water for Injection q.s. q.s. q.s.

The above mentioned compositions of paclitaxel and albumin nanosuspension were prepared in accordance with the present invention in the following manner:
a) preparing aqueous phase by dissolving human albumin in water
b) preparing non-aqueous phase by dissolving paclitaxel in organic solvent
c) preparation of crude emulsion of (a) & (b)
d) preparation of nanosuspension of (c)
e) dilution of nanosuspension of (d) with water
f) removal of solvent
g) aseptic filtration
h) filling followed by
i) lyophilization

Table 2 discloses the effect of dilution with water (step-e) on particle size of paclitaxel-albumin nanoparticles. The particle size was measured using Zetasizer apparatus.

Table 2: Effect of dilution on particle size of paclitaxel-albumin nanoparticles
Particle size (nm)
T0hr T1hr T24hrs T3months
(40 oC/75% RH ) T6months
(40 oC/75% RH )
Undiluted bulk 100-230 230-500 500-700 NA NA
Diluted bulk 100-230 100-230 100-230 100-230 100-230

Table 3: Stability data of reconstituted paclitaxel and albumin nanosuspension
Test Initial 2 to 8ºC Room temperature
8 hrs 24 hrs 8 hrs 24 hrs
Sedimentation No visible sediment No visible sediment No visible sediment
Particle Size (nm) 100-230 100-230 100-230 100-230 100-230
Assay of Paclitaxel (%) Not less than 90% and Not more than 110 % of the labeled amount of Paclitaxel

,CLAIMS:1. A process for preparation of paclitaxel and albumin nanosuspension comprising:
a) preparing aqueous phase by dissolving human albumin in water
b) preparing non-aqueous phase by dissolving paclitaxel in organic solvent
c) preparation of crude emulsion of (a) & (b)
d) preparation of nanosuspension of (c)
e) dilution of nanosuspension of (d) with water
f) removal of solvent
g) aseptic filtration
h) filling and
i) lyophilization
2. The process as claimed in claim 1, wherein the organic solvents for step (b) are selected from the group comprising of aliphatic, cycloaliphatic, or aromatic hydrocarbons having 4-30 carbon atoms such as n-dodecane, n-decane, n-hexane, cyclohexane, toluene, benzene, and mixtures thereof, aliphatic or aromatic alcohols having 2-30 carbon atoms such as ethanol, octanol and mixtures thereof, alkyl, aryl, or cyclic ethers having 2-30 carbon atoms such as diethyl ether, tetrahydrofuran, and mixtures thereof, alkyl or aryl halides having 1-30 carbon atoms selected from the group of CHCl3, CH2Cl2,CH3ClCH2Cl2, CH2Cl-CH2Cl, and mixtures thereof, aliphatic or aromatic esters having 2-30 carbon atoms such as ethyl acetate, methyl acetate, ethyl caprylate (octanoate) and mixtures thereof , ketones having 3-30 carbon atoms such as acetone, methyl ethyl ketone and mixtures thereof , polyalkylene glycols such as polyethylene glycol, and mixtures thereof.
3. The process as claimed in claim 1, wherein crude emulsion was prepared by using homogenization or mixing or sonication method.
4. The process as claimed in claim 1, wherein nanosuspension in step (d), was prepared by using high pressure homogenizer or ultrasonication or extrusion or high energy wet milling technique.
5. The process as claimed in claim 4, wherein step (d) was performed till particle size of paclitaxel- albumin nanoparticle not more than 1000 nm is achieved.
6. The process as claimed in claim 5, wherein step (d) was performed till particle size of paclitaxel- albumin nanoparticle is not more than 230 nm.
7. The process as claimed in claim 1, wherein the nanosuspension obtained after step (d) was diluted with water.
8. The process as claimed in claim 7, wherein the amount of water added for dilution was 2 to 10 times the quantity of nanosuspension bulk of step (d).
9. The process as claimed in claim 1, wherein the solvent was removed by using tangential flow filtration, or wiped film evaporator or thin film evaporator or rotary evaporator or combinations thereof.
10. The process of claim 1, wherein the nanosuspension of step (f) is aseptically filtered, filled in vials and lyophilized.