FORM 2
THE PATENTS ACT, 1970
(39 of 1970)
&
The Patent Rules, 2003
COMPLETE SPECIFICATION (See section 10 and rule 13)
STABLE PHARMACEUTICAL COMPOSITION
SUN PHARMA ADVANCED RESEARCH COMPANY LTD.
A company incorporated under the laws of India having their office at 17/B, MAHAL INDUSTRIAL ESTATE, MAHAKALI CAVES ROAD, ANDHERI (E), MUMBAI-400093, MAHARASHTRA, INDIA.
The following specification particularly describes the invention and the manner in which it is to be performed.

FIELD OF THE INVENTION
The present invention relates to a novel pharmaceutical composition in the form of a solution and a nanodispersion of a water insoluble active ingredient.
BACKGROUND OF THE INVENTION
It is believed that almost 40 % of therapeutically active compounds identified through combinatorial screening programs are poorly soluble in water. Such water insoluble drugs are commnly formulated into liquid formulations as suspension and the suspension formulations thereof are associated with problems such as physical instability, clarity, particle size change, crystal form instability, poor bioavailability and the like. The present invention relates to pharmaceutical composition in the form of a solution. In an attempt to develop such pharmaceutical composition in the form of a solution, inventors have disclosed in their earlier PCT publication number WO2010/146606 a nanodispersion comprising nanoparticles of mean size less than 300 nm dispersed in a vehicle comprising water miscible solvent and water, said nanoparticles comprising one or more drugs, a polymer and a surfactant comprising a mixture of fatty acids or its salts and sterol or its derivatives or its salts.
While attempting to further stabilize a pharmaceutical composition in the form of a solution that is stable on storage and/or after reconstitution, the inventors surprisingly and unexpectedly observed that a sufficient amount of the close structural analog, derivative, pro-drug or metabolite of the active ingredient can provide improved stability to the pharmaceutical composition in the form of a solution. The pharmaceutical composition in the form of a solution is designed to form nanoparticles in vivo when exposed to the appropriate biological fluid or reconstituted with an infusion fluid such as dextrose, saline or a buffer. The pharmaceutical composition in the form of a solution is stable physically and chemically in the form of pharmaceutical composition in the form of a solution as well as nanodispersion. It was indeed surprising to find that the addition of a sufficient amount of close structural analog of the water insoluble active ingredient to the water insoluble active ingredient provides a physically stable pharmaceutical composition in the form of a solution which is stable upon reconstitution with aqueous vehicle.

OBJECTS OF THE INVENTION
It is an object of the present invention to provide a pharmaceutical composition in the
form of a solution of a water insoluble active ingredient that forms a stable
nanodispersion upon reconstitution with aqueous vehicle.
Further it is the object to provide a pharmaceutical composition in the form of a solution
that is easy to manufacture, scale-up and use for administration.
It is an object of the present invention to provide a pharmaceutical composition in the
form of a solution of a water insoluble active ingredient that provides safety and
satisfactory efficacy.
SUMMARY OF THE PRESENT INVENTION
The present invention provides a pharmaceutical composition in the form of a solution comprising water insoluble active ingredient and a close structural analog of the water insoluble active ingredient, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the water insoluble active ingredient is present in therapeutically effective amounts and the close structural analog is present in sufficient amounts such that when the composition is added to an aqueous vehicle it forms a nanodispersion suitable for intravenous infusion, wherein said nanodispersion is stable for longer periods as compared to a dispersion formed from an identical composition but devoid of the close structural analog of the water insoluble active ingredient.
In another aspect, the present invention provides a pharmaceutical composition in the form of a solution comprising water insoluble active ingredient and a close structural analog of the water insoluble active ingredient, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the water insoluble active ingredient is present in therapeutically effective amounts and the close structural analog is present in sufficient amounts such that when the composition is orally administered, it forms a nanodispersion wherein the particle size of the water insoluble inactive ingredient is maintained below 1000 nanometers for a sufficient time during which absorption through the mucosa takes place.

BRIEF DESCRIPTION OF FIGURE
Figure 1 shows the results of the in-vivo efficacy study of the pharmaceutical composition of the present invention in athymic nude mice bearing Human Mammary Carcinoma (MX-1) xenografts. The graph shows tumor volume data plotted against number of days post treatment
Figure 2 shows the results of the in-vivo efficacy study of the pharmaceutical composition of the present invention in athymic nude mice bearing Human Colorectal Carcinoma (HT-29) xenografts. The graph shows tumor volume data plotted against number of days post treatment
Figure 3 shows the results of the in-vivo efficacy study of the pharmaceutical composition of the present invention in athymic nude mice bearing Human Prostate Carcinoma (PC-3) xenografts. The graph shows tumor volume data plotted against number of days post treatment
DETAILED DESCRIPTION OF THE INVENTION
The term 'nanodispersion' as defined herein means that the pharmaceutical composition in the form of a solution or solution comprising water insoluble active ingredient, when dispersed in an aqueous vehicle, the active ingredients are in the form of nanoparticles having a particle size less than 2000 nm, preferably, less than 500 nm, preferably the particles are colloidal in nature. The dispersed particles may also be in the form of micelles, compound micelles, vesicles, mesophases, nanostructured liquid crystalline phases, ribbons, nano rods, helices, cubosomes, hexosomes or like.
According to the present invention, the term 'physically stable' as used herein, means the pharmaceutical composition in the form of solution of the present invention does not show any sign of crystallization or precipitation. When the pharmaceutical composition in the form of a solution is diluted in aqueous vehicle, it forms a nanodispersion that is suitable for intravenous infusion. The nanodispersion is said to be physically stable for longer periods when there are no signs of changes in the appearance such as change from translucent to hazy or signs of settlement of particles or signs of turbidity or the particle size changing to more than 1000 nm. Any one of these changes in the nanodispersion is

considered as signs of instability and the nanodispersion is then said to be 'unstable.' The nanodispersion is said to be stable for certain duration of time during which none of the above mentioned changes are observed. In one embodiment, the pharmaceutical composition in the form of a solution is found to be physically stable when upon dispersion in the aqueous vehicle, the nanodispersion so formed, remains stable for at least 30 minutes, preferably 2 hours and most preferably 8 hours or upto 24 hours. During this time the nanodispersion may be administered via parenteral route at a rate suitable for intravenous administration as an infusion.
The term 'water insoluble' as used herein means the active ingredient are sparingly soluble or are less than sparingly soluble, including practically insoluble compounds. Sparingly soluble compounds have solubility ranging from 10 to 33 mg/ml, slightly soluble compounds from 1-10 mg/ml and very slightly soluble compounds from 0.1 to 1 mg/ml. Active ingredients with solubility below 1 mg/ml are classified as practically insoluble. Examples of some of the water insoluble active ingredients are fenofibrate, taxane derivatives like paclitaxel, docetaxel, SN-38 (active moiety of irinotecan), tacrolimus, sirolimus, temsirolimus, cyclosporine, pirenoxine, nepafenac, brinzolamide, temozolamide, etoposide and like.
The 'close structural analog' of the water insoluble active ingredient includes any chemical compound having some structural similarity to the chemical structure of the water insoluble active ingredient. The close structural analog may be any one such as the derivative, ester, prodrug or a metabolite of the same active ingredient. For example, a taxane compound such as paclitaxel can be used to stabilize another taxane, such as docetaxel. Similarly, one camptothecin compound can be used to stabilize another camptothecin compound. Without wishing to be bound by the theory, the analogs are capable of hydrophobically interacting with the water insoluble active ingredient due to the similar back bone or a basic chemical structure and in addition they may have a charged or a hydrogen bonding group that provides interfacial stabilization of the nanoparticles in aqueous medium.


.3H2O
Docetaxel: (2R,3S)-N-carboxy-3-phenylisoserine, N-tert-butyl ester, 13-ester with 5, 20-
epoxy-1,2,4, 7,10,13-hexahydroxytax-l l-en-9-one 4-acetate 2-benzoate, trihydrate.

Close structural analog of docetaxel: Paclitaxel: (2α,4α,5β,7β,10β,13α)-4,10-bis(acetyloxy)-13-{[(2R,3S)- 3-(benzoylamino)-2-hydroxy-3-phenylpropanoyl]oxy}- 1,7-dihydroxy-9-oxo-5,20-epoxytax-l l-en-2-yI benzoate

SN3 8: 7-Ethyl-10-Hydroxy~20(S)-Camptothecin

Close structural analog of SN-38 : Irinotecan: (S)-4,11-diethyl-3,4,12,14-tetrahydro-4-hydroxy-3,14-dioxolH-pyrano[3',4' :6,7]-indolizino[l,2-b]quinolin-9-yl-[1,4'bipiperidine]-l '-carboxylate

Fenofibrare: Propan-2-yl 2-{4-[(4-chlorophenyi)carbonyl]phenoxy} -2-methylpropanoate


Close structural analog of fenofibrate: Fenofibric acid: 2-[4'-(p-Chlorobenzoyl) phenoxy]-2-methylpropionic acid
According to the present invention, the close structural analog present in the pharmaceutical composition is present in 'sufficient amounts' such that when the composition is added to an aqueous vehicle, it forms a nanodispersion suitable for intravenous infusion wherein said nanodispersion is stable for longer periods as compared to a dispersion formed from an identical composition but devoid of the close structural analog of the water insoluble active ingredient. In another aspect, 'sufficient amounts' of the close structural analog present in the pharmaceutical composition is such that when the composition is orally administered, it forms a nanodispersion wherein the water insoluble inactive ingredient remains as particles having particle size less than 1000 nm, preferably, less than 800 nm and still preferably about 500 nm, for time period sufficient to allow its absorption through the intestinal mucosa.
When the pharmaceutical composition of the present invention is added to the aqueous vehicle, if the dispersion so formed shows any one of the changes such as change in the appearance or change in particle size or signs of precipitation or turbidity or haziness in the nanodispersion, the nanodispersion is considered to be 'unstable.' The nanodispersion is said to be stable for certain duration of time in which time none of the above mentioned changes are observed. In one embodiment, the pharmaceutical composition in the form of a solution is found to be physically stable when upon dispersion in the aqueous vehicle, the nanodispersion so formed, remains stable for at least 30 minutes, preferably 2 hours and most preferably 8 hours or upto 24 hours. During this time, the

nanodispersion may be administered via parenteral route at a rate suitable for intravenous administration as an infusion. In another aspect, the pharmaceutical composition may be administered orally such that the nanodispersion is formed and the active ingredient remains in nanoparticles wherein effective absorption of the water insoluble active ingredient takes place. It is found that the amount of the close structural analog of a water insoluble active ingredient varies from one active ingredient to another. It is believed that this depends upon the nature and the chemistry of the water insoluble active ingredient. For example, for water insoluble active ingredient like SN-38, when irinotecan was used to stabilize, it was found that the pharmaceutical composition in the form of a solution and nanodispersion was stable in terms of the physical stability when irinotecan was present in an amount of more than about 23% w/w of the total amount of the water insoluble active ingredient, SN-38. In other words, a molal ratio of SN-38 and close structural analog of SN-38, irinotecan of less than 7, preferably about 5, most preferably about 4, was found to provide a stable pharmaceutical composition in the form of a solution of the present invention. In certain embodiments, analog of the SN-38 other than irinotecan may be used in the pharmaceutical composition in the form of a solution of the present invention. Examples of the other analogs that may be used include, but are not limited to, camptothecin, 9-aminocamptotothecin, 9-nitrocamptothecin, topotecan, gimatecan, lurtotecan, rubitecan, SN-38G (10-O-glucoronyl-SN-38) and other camptothecin analogs, quinoline and its analog and their mixtures. In case of other embodiments where pharmaceutical composition in the form of a solution comprising docetaxel is stabilized with a structural close analog, a molal ratio of docetaxel to close structural analog, paclitaxel of less than 25, preferably about 20, mostly preferably about 10 was found to provide stable pharmaceutical composition in the form of a solution. In another such similar embodiments of the present invention, docetaxel was formulated as a pharmaceutical composition in the form of a solution comprising its structural analog, selected from the group consisting of paclitaxel, taxol derivative, partial analogs, intermediates like threo-2-(l-Ethoxyethoxy)-3-tert-butyloxy carbonylamino-3-phenylpropanoic acid, 10-deacetyl baccatin and baccatin analog. A typical example contains docetaxel, paclitaxel, polyvinylpyrrolidone, sodium cholesterol sulfate, caprylic acid and water miscible solvents like ethanol and polyethylene glycol. The

pharmaceutical composition in the form of a solution when reconstituted with dextrose solution was converted to nanodispersion. The nanoparticles docetaxel and paclitaxel as a stabilizer were found to be in the form of spherical micelles. In one embodiment, pharmaceutical composition in the form of a solution of the fenofibrate is stabilized by inclusion of fenofibric acid which is close structural analog. In one particular embodiment, the molal ratio of fenofibrate to fenofibric acid of about 9 was found to provide a stable pharmaceutical composition in the form of a solution which also provided a stable nanodispersion when diluted with aqueous vehicle. Without wishing to be bound by any theory, it may be said that the amount of the close structural analog in the pharmaceutical composition varies depending upon the water insoluble active ingredient.
The present invention provides a pharmaceutical composition in the form of a solution comprising water insoluble therapeutically active ingredient and a close structural analog of the water insoluble active ingredient, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the water insoluble active ingredient is present in therapeutically effective amounts and the close structural analog is present in sufficient amounts such that when the composition is added to an aqueous vehicle it forms a nanodispersion suitable for intravenous infusion wherein said nanodispersion is stable for longer periods as compared to a dispersion formed from an identical composition but devoid of the close structural analog of the water insoluble active ingredient.
According to one preferred embodiment, the present invention provides a pharmaceutical composition in the form of a solution comprising a water insoluble active ingredient and its close structural analog and one or more water miscible solvents and one or more pharmaceutically acceptable excipients. In one preferred embodiment, the pharmaceutical acceptable excipients include, but are not limited to, water soluble polymers, one or more surfactants and mixtures thereof.

The water miscible solvent used in the pharmaceutical composition of the present
invention is one in which the water insoluble active ingredient is relatively soluble and
which is miscible with water or aqueous solvents. Examples of such solvents include, but
are not limited to: alcohols such as ethanol, n-propanol, isopropanol; glycols such as
ethylene glycol, propylene glycol, butylene glycol and its derivatives; polyethylene
glycols like PEG 400 or PEG 3350; polyethylene glycol esters and ethers such as
polyethylene 15 glycol sorbitans, polyethylene glycol monoalkyl ethers; polypropylene
glycol and its derivatives such as PPG-10 butanediol, PPG-10 methyl glucose ether, PPG-
20 methyl glucose ether, PPG- 15 stearyl ether; glycerol; glycofurol; dimethylsulfoxide
(DMSO); dimethylacetamide; dimethylformamide; 1,4-dioxane, dimethylisosorbide,
dimethylnicotinamide and the like and mixtures thereof. In one embodiment of the
present invention, the non-aqueous solvent may be selected from alcohols, polyethylene
glycols and/or mixtures thereof. In preferred embodiment of the present invention, a
mixture of ethanol and PEG (polyethylene glycol) is used as the non-aqueous solvent.
Ethanol is used in the pharmaceutical composition in the form of a solution of the present
invention in an amount ranging from about 0.001% w/v to about 50% w/v, more
preferably from about 0.01 % w/v to about 20% w/v and most preferably from about 1%
w/v to about 10% w/v. Polyethylene glycols which are used preferably, include
Polyethylene glycol 400 and Polyethylene glycol 3350. In certain embodiments, PEG-
400 is used in an amount ranging from about 20 % w/v to about 95.0% w/v, more
preferably from about 60% w/v to about 90% w/v and most preferably from about 75 %
w/v to about 85 % w/v of the pharmaceutical composition in the form of a solution. In
some embodiments, a combination of two or more water miscible solvents may be used
for example; a combination of polyethylene glycol, polyvinyl pyrollidone and water
miscible solvent such as dimethyl sulfoxide may be used. The amount of the organic
solvents such as ethanol, dimethyl sulfoxide may vary from about 2 % to about 50 %
w/w, preferably 10 % to 40 %. However, depending upon the water insoluble active
ingredient, the range may vary. For instance, in case of SN-38, the amount of the
dimethylsulfoxide in the pharmaceutical composition in the form of a solution is about 10
% w/w, whereas in case of fenofibrate the amount of organic solvents such as a
combination of ethanol and dimethylsulfoxide in the pharmaceutical composition in the

form of a solution is about 40 % w/w and in case of drugs like docetaxel, the amount of organic solvent in the pharmaceutical composition in the form of a solution is about 10 % w/w. It may be noted that the choice of the type of water miscible solvents will depend upon the water insoluble active ingredient that is to be formulated as nanodispersion as well as upon the mode of its administration such as parenteral or oral.
The water soluble polymer(s) that may be used in the pharmaceutical composition of the
present invention are selected from a group consisting of tertiary amide polymers such as
polyvinylpyrrolidone and the like (PVP-PEG conjugate); polyamino acids such as
polyglutamic acid and poly-1-lysine; polysaccharide containing glycos-aminoglycans
such as hyaluronan, heparin sulphate and chondroitin sulphate; natural polymers such as
gelatin, chitosan, human serum albumin; and the like; polyanhydrides such as poly
sebacic acid: polyamides such as polyglutamates and other biocompatible and
biodegradable polymers and/or their derivatives or copolymers or any other such
polymers known to one skilled in the art and mixtures thereof According to one
embodiment of the present invention, the polymer used is polyglutamic acid sodium salt.
According to another embodiment of the present invention, the polymer used is sodium
hyaluronate. According to preferred embodiment of the present invention, the polymer
used is polyvinylpyrrolidone. Polyvinylpyrrolidone is a tertiary amide polymer having
linearly arranged monomer units of l-vinyl-2-pyrrolidone, hereinafter designated PVP,
and also known as Povidone. It is commercially available as a series of products having
mean molecular weights ranging from about 10,000 to about 700,000. The various
products are marketed according to average molecular weights designated K-values; e.g.
GAF Corporation supplies PVP having the following K-values: K 15 is about 10,000; K
30 is about 40,000; K 60 is about 160,000 and K 90 is about 360,000. The pharmaceutical
composition in the form of a solution of this invention may contain various grades of
polyvinylpyrrolidone, i.e. for example, PVP K-12, K-17, K-30, K-60 and K-90. The
polyvinylpyrrolidone ingredient may be present as one specific grade or as a combination
of two or more grades. According to one embodiment of the present invention, the
polymer may be used in the pharmaceutical composition in the form of a solution in an
amount ranging from about 0.001% w/w to about 50% w/w. The polymer is preferably

used in an amount ranging from about 1% w/w to about 40% wAv. Most preferably, it is used in an amount ranging from about 2 % w/w to about 20% w/w.
The pharmaceutical composition of the present invention may comprise a
pharmaceutically acceptable excipient such as water soluble polymers, surfactants or
mixtures thereof. . The term surfactant is a blend of "surface active agent". Surfactants
are molecules, which comprises a water-soluble (hydrophilic) and a lipid-soluble
(lipophilic) part. The surfactants that may be used in the pharmaceutical composition of
the present invention may be selected from nonionic, ionic, anionic, cationic, and
zwitterionic surfactants. The surfactants that may be used in the pharmaceutical
composition of the present invention may be selected from cetostearyl alcohol,
cetomacrogol emulsifying wax, sorbitan esters, polyoxyethylene alkyl ethers (e.g.,
macrogol ethers such as cetomacrogol 1000), polyoxyethylene castor oil derivatives,
polyoxyethylene sorbitan fatty acid esters (e.g., the commercially available Tweens®
such as e.g., Tween® 20 and Tween® 80; polyethylene glycols (e.g., Carbowaxes 3550®
and 934'), polyoxyethylene stearates, phosphates, carboxymethylcellulose calcium.
carboxymethylcellulose sodium, methylcellulose, hydroxyethylcellulose, hypromellose
phthalate, noncrystalline cellulose, triethanolamine, polyvinyl alcohol (PVA), 4-(l, 1,3,3-
tetramethylbutyl)-phenol polymer with ethylene oxide and formaldehyde (also known as
tyloxapol, superione, and triton), poloxamers (e.g., Pluronics® F68 and F108, which are
block copolymers of ethylene oxide and propylene oxide); poloxamines (e.g., Tetronic
908®, also known as Poloxamine 908®, which is a tetrafunctional block copolymer
derived from sequential addition of propylene oxide and ethylene oxide to
ethylenediamine-BASF); Tetronic 1508® (T-1508) (BASF), Tritons X-200®, which is an
alkyl aryl polyether sulfonate; mixture of sucrose stearate and sucrose distearate, PEG-
phospholipid, PEG-cholesterol, PEG-cholesterol derivative, PEG-vitamin A. PEG-
vitamin E, lysozyme, random copolymers of vinyl pyrrolidone and vinyl acetate, and the
like. In one embodiment, the surfactants that are used in the pharmaceutical composition
of the present invention may be selected from fatty acids or its salts, sterol or its
derivatives including salts and mixtures thereof. The term fatty acids includes aliphatic
(saturated or unsaturated) monocarboxylic acids derived from or contained in esterified

form, in an animal or vegetable fat, oil or wax. Examples of fatty acids or its salts that
may be used in the pharmaceutical composition in the form of a solution of the present
invention include but are not limited to fatty acids or its salts having 'n' number of
carbon atoms wherein 'n' ranges from about 4 to about 28. The fatty acid may be a
saturated fatty acid or an unsaturated fatty acid, and their salt and combinations thereof.
The saturated fatty acid and its salts may be selected from butyric acid, caproic acid,
caprylic acid, capric acid, lauric acid, myristic acid, palmitic acid, stearic acid, arachidic
acid, behenic acid, sodium caprylate, sodium laurate, sodium myristate, sodium palmitate
and the like and/or mixtures thereof. The unsaturated fatty acid and its salts may be
selected from myristoleic acid, palmitoleic acid, oleic acid, linoleic acid, alpha linolenic
acid, arachidonic acid, eicosapentaenoic acid, erucic acid, docosahexaenoic acid, sodium
oleate, sodium arachidonate and the like and/or mixtures thereof. Examples of sterol or
its derivative or its salts that may be used in the pharmaceutical composition in the form
of a solution of the present invention may be selected form cholesterol, phytosterols,
ergosterol, bile acids and their derivatives, salts and mixtures thereof. Cholesterol its
derivatives and salts include cholesteryl sulfate, cholesterol acetate, cholesterol
chloroacetate, cholesterol benzoate, cholesterol myristate, cholesterol hemisuccinate,
cholesterol cinnamate, cholesterol crotanoate, cholesterol butyrate, cholesterol
heptanoate, cholesterol hexanoate, cholesterol octanoate, cholesterol nonanoate,
cholesterol decanoate, cholesterol oleate, cholesterol propionate, cholesterol valerate,
dicholesteryl carbonate, and the like. Phytosterols that may be used in the pharmaceutical
composition in the form of a solution of the present invention include sitosterol,
campesterol, stigmasterol, brassicasterol and its derivatives, salts and mixture thereof. For
example, Phytosterols marketed by Sigma, U.S.A. containing bsitosterol, campesterol
and dihydrobrassicasterol. Bile acids include cholic acid, chenodeoxycholic acid,
deoxycholic acid, glycocholic acid, taurocholic acid, ursodeoxycholic acid and its
derivatives, salts and mixture thereof. According to one embodiment of the present
invention, the surfactant used is selected from saturated fatty acid and cholesterol salt
and/or mixtures thereof. According to preferred embodiment of the present invention, the
surfactant used is selected from caprylic acid and cholesteryl sulphate and/or mixtures
thereof. In one embodiment, a combination of cholesteryl sulphate and caprylic acid are

used as surfactant in the pharmaceutical composition in the form of a solution in the range of about 0.10 % to about 5 %, preferably about 2 % w/w. It is also possible to possible to use other surfactants.
In one specific embodiment of the present invention, the surfactant used in the pharmaceutical composition is a mixture of caprylic acid and cholesteryl sulfate. Caprylic acid that may be used in the embodiments is in an amount ranging from about 0.001 % w/v to about 5.0 % w/v, more preferably from about 0.01 %w/v to about 1.0 % w/v and most preferably from about 0.01 %w/v to about 0.5% w/v. Cholesteryl sulfate is used in the embodiments of the present invention in an amount ranging from about 0.001 % w/v to about 5.0% w/v, more preferably from about 0.01 %w/v to about 1.0 %w/v and most preferably from about 0.01 % w/v to about 0.5 % w/v. According to another embodiment of the present invention, the surfactant used is selected from unsaturated fatty acid and cholesterol salt and/or mixtures thereof. According to preferred embodiment, the surfactant used is selected from oleic acid or caprylic acid and cholesteryl sulphate and/or mixtures thereof. According to another embodiment of the present invention, the surfactant used is selected from saturated fatty acid and bile acid or bile salt and/or mixtures thereof. According to preferred embodiment, the surfactant used is selected from caprylic acid and sodium glycocholate or ursodeoxycholic acid and/or mixtures thereof.
In one embodiment of the present invention, the water soluble polymer used in the pharmaceutical composition is povidone. Povidone is used in the concentration ranging from 0.01 % w/w to 20 % w/w more preferable from 0.1% w/w to 10 % w/w and most preferred is 7.5 % w/w of the pharmaceutical composition in the form of a solution. Upon dilution with the aqueous vehicle, the concentration of the polymer in the nanodispersion ranges from 0.001 % to 1.0 % w/w. In one embodiment of the present invention, the solvents used in the pharmaceutical composition in the form of a solution are ethanol and PEG-400. Ethanol is used in the concentration ranging from 5 % to 95 % more preferable from 1 to 20% and most preferred is 10 % in the pharmaceutical composition in the form of a solution. After dilution the concentration will be decreased and it may range from

0.001% to 2.0%. PEG-400 is used in the concentration ranging from 5 % to 95 % more preferable from 75 % to 85 % and most preferred is 80 % in the pharmaceutical composition in the form of a solution. After dilution the concentration will be decreased and it may range from 1 % to 20 %.
In one embodiment, the pharmaceutical composition in the form of a solution comprises SN-38 (7-ethyl 10-hydroxy camptothecin) as the water insoluble active ingredient and irinotecan as the close structural analog. It was found that the pharmaceutical composition in the form of a solution was stable in terms of the physical stability when irinotecan was present in amount of more than about 23 % of the total amount of the water insoluble active ingredient, SN-38. In other words, a molal ratio of SN-38 and close structural analog of SN-38, irinotecan of less than 7, preferably about 5, mostly preferably about 4, was found to provide a stable pharmaceutical composition in the form of a solution of the present invention. In one particular embodiment, the present invention provides a pharmaceutical composition in the form of a solution comprises SN-38 and irinotecan and one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the SN-38 is present in therapeutically effective amounts and irinotecan is present in sufficient amounts such that when the composition is added to an aqueous vehicle it forms a nanodispersion suitable for intravenous infusion wherein said nanodispersion is stable for longer periods as compared to a dispersion formed from an identical composition but devoid of the close structural analog of the water insoluble active ingredient. The pharmaceutically acceptable excipients include a water soluble polymers or surfactants or mixtures thereof. In one preferred embodiment, the water soluble polymer used is polyvinyl pyrrolidone and the surfactant used is a mixture of fatty acid and a cholesterol derivative. One particular embodiment provides a pharmaceutical composition in the form of a solution comprising SN38, irinotecan, polyvinylpyrrolidone, sodium cholesteryl sulfate, caprylic acid and water miscible solvents like dimethyl sulfoxide, ethanol and polyethylene glycol in the mixture of two or more. The pharmaceutical composition in the form of a solution when reconstituted with dextrose solution was found to provide a stable nanodispersion and the nanoparticles were found to be present in the form of helical ribbons.

In one aspect, the present invention provides its applicability to improve the oral bioavailability of a water insoluble active ingredient, for example., fenofibrate. Fenofibrate is known to be a notorious because of the problems associated with the oral bioavailability. In this aspect, the present invention provides a pharmaceutical composition in the form of a solution comprising water insoluble active ingredient and a close structural analog of the water insoluble active ingredient, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the water insoluble active ingredient is present in therapeutically effective amounts and the close structural analog is present in sufficient amounts such that when the composition is orally administered, it forms a nanodispersion wherein the particle size of the water insoluble inactive ingredient is maintained below 1000 nanometers for a sufficient time during which absorption through the mucosa takes place. In one particular embodiment related to this aspect, it was found that a water insoluble active ingredient, fenofibrate could be formulated effectively to improve its oral bioavailability. The pharmaceutical composition of fenofibrate according to the present invention was prepared as a physically stable composition in the form of a solution with the help of addition of sufficient amounts of its close structural analog, fenofibric acid. In one specific embodiment, a molal ratio of fenofibrate to fenofibric acid of about 9 was found to provide a stable pharmaceutical composition in the form of a solution of the present invention. Fenofibrate is formulated as a solution wherein the pharmaceutical composition comprises it close structural analog such as fenofibric acid, 4-chlorobenzophenone analogs or benzophenone analogs. In one particular embodiment, fenofibric acid is used to stabilize fenofibrate. The amount of fenofibric acid used ranges from about 0.001% to 50 % w/w, preferably 1 to 20 % w/w of the pharmaceutical composition in the form of a solution or about 0.1 to 10 % w/w of the nanodispersion. For water insoluble active ingredient like fenofibrate, it was found that the pharmaceutical composition in the form of a solution was stable for about 1 hour to 2 hours, after which there were signs of aggregation. The use of fenofibric acid increases the solubility of fenofibrate in the pharmaceutical composition in the form of a solution. The nanodispersion of the present invention comprises nanoparticles dispersed in a vehicle comprising non-aqueous solvent and water. The minimum approved strength of

fenofibric acid is 35 mg oral. According to the present invention, preferably the fenofibrate dose will not exceed to a dose which will deliver 35 mg of fenofibric acid in the said pharmaceutical composition in the form of a solution.
The present invention can be said to overcome the problems of poor bioavailability of fenofibrate by orally administering the pharmaceutical composition in the form of a solution comprising fenofibrate and its close structural analog such as for example, fenofibric acid, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein fenofibrate is present in therapeutically effective amounts and the close structural analog such as fenofibric acid, is present in sufficient amounts such that when the composition is orally administered, it forms a nanodispersion wherein the fenofibrate remains as particles having size less than 1000 nm for time sufficient to allow its absorption through the intestinal mucosa takes place. Thus the present invention can be said to provide a method of improving the oral bioavailability of fenofibrate. In these embodiments, the pharmaceutical composition in the form of a solution may be filled into soft gelatin capsules, or the pharmaceutical composition in the form of a solution may be sprayed on an inert carrier and then the carrier may be converted into any other solid dosage form like powder filled into hard gelatin capsules, compressed dosage form like tablets, caplets. When the pharmaceutical composition in the form of a solution in the form of solution is filled into soft gelatin capsules, the soft gelatin capsules upon oral administration, would convert the pharmaceutical composition in the form of a solution into nanodispersion. In one embodiment, fenofibrate, an orally administered active agent is formulated into the pharmaceutical composition in the form of a solution of the present invention by incorporating fenofibric acid to stabilize fenofibrate.
The pharmaceutical composition in the form of a solution of the present invention is prepared by simple process. The process includes steps of stirring a mixture of water insoluble active ingredient, sufficient amounts of structural analog or prodrug or salt or its derivative, one or more polymer(s) and/ or surfactant(s) such as fatty acids or its salts, sterol or its derivatives including salts thereof is in non-aqueous solvent such as ethanol

and/or PEG) with or without the application of heat treatment to obtain a pharmaceutical composition in the form of a solution of the drug. The pharmaceutical composition in the form of a solution so obtained is aseptically filtered through a 0.2um membrane filter to obtain a pharmaceutical composition in the form of a solution of the present invention. To this pharmaceutical composition in the form of a solution, at the time of administration, an aqueous liquid vehicle (5% w/v dextrose solution) may be added slowly and the mixture is shaken/ agitated, to obtain a nanodispersion. Generally, the pharmaceutical composition in the form of a solution of the present invention is diluted with a vehicle which comprises water as sterile water for injection or about 5% to about 10.0% w/v dextrose solution or about 0.45% to about 0.9% w/v normal saline solution or a combination of 2.5% and 0.9% sodium chloride solution or any other pharmaceutically acceptable intravenous aqueous liquid vehicle and mixtures thereof. In preferred embodiments, 5% w/v and 10% w/v dextrose solution is used as the aqueous liquid vehicle. Vehicles are particularly important as it helps to minimize the effect of drug, especially an anticancer drug on site of infusion. Solutions of dextran, albumin, hetastarch, hyaluronate and other biopolymers that are generally to improve the physical stability of the nanodispersion obtained upon admixture of the pharmaceutical composition in the form of a solution.
The present invention is said to provide a pharmaceutical composition provides a pharmaceutical composition in the form of a solution comprising water insoluble active ingredient and a close structural analog of the water insoluble active ingredient, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the water insoluble active ingredient is present in therapeutically effective amounts and the close structural analog is present in sufficient amounts such that when the composition is added to an aqueous vehicle it forms a nanodispersion suitable for intravenous infusion wherein said nanodispersion is stable for longer periods as compared to a dispersion formed from an identical composition but devoid of the close structural analog of the water insoluble active ingredient. The improvement in the time period in which the nanodispersion remains stable provides advantages especially when the composition is administered to patients as infusion drips. Since the size of the

dispersed particles remains constant and there is no precipitation during administration, the present invention can be said to provide a method of administering a water insoluble active ingredient, wherein the composition in the form of a solution when added to aqueous vehicle, it forms a nanodispersion suitable for intravenous infusion. Since the nanodispersion remains stable for a considerable period of time, the infusion may be kept for repeated dosing instead of making a fresh infusion at the time of administration in situations where the nanodispersion is not stable for longer period of times. Thus, the present invention is overcome the problem associated with the precipitation of a water insoluble active ingredient. It is always advantageous to have a nanodispersion that is made by a very simple, quick, less time consuming reconstitution procedure. The present invention thus provides application for administering water insoluble drugs as intravenous infusions, e.g. reconstitution done next to the patient on bed under medical supervision from the pharmaceutical composition which is in the form of a solution.
According to one embodiment of the present invention, the pharmaceutical composition in the form of a solution was subjected to safety studies in Balb/c nude mice by observing the mortality after intravenous administration of the pharmaceutical composition in the form of a solution of the present invention. The maximum tolerated dose of SN-38 in the pharmaceutical pre-concentrate of the present invention was found to be more than 7.5 mg/kg and less than 10 mg/kg following intravenous injection. Further, the present invention can be said to provide a method of treating a disease by administration a pharmaceutical composition in the form of a solution or the diluted pharmaceutical composition in the form of a solution i.e. the nanodispersion of the water insoluble active ingredient such as SN-38 or docetaxel to human or animals for the prevention. amelioration and/or cure of cell proliferative diseases, such as cancer. The pharmaceutical composition in the form of a solution of the present invention can be said to be useful in treating human lymphoma, ovarian, breast, prostrate, colon, lung cancers as well as multiple sclerosis.
The pharmaceutical composition in the form of a solution of the present invention can be provided as a kit having two containers, the first container comprising the pharmaceutical

composition in the form of a solution of one or more water insoluble active ingredient, sufficient amounts of close structural analog of the same water insoluble active ingredient, a polymer and a surfactant selected from fatty acids or its salts, sterol or its derivatives including salts and mixtures thereof in a water miscible solvent, and a second container comprising an aqueous liquid vehicle, for example infusion fluid, such that on addition of contents of second container to the contents of the first container or vice versa, with mild agitation or shaking, results in the formation of nanodispersion of the present invention and is suitable for intravenous administration as an infusion.
The pharmaceutical composition in the form of a solution of the present invention can also provides a kit having two containers, the first container comprising a lyophilized form of the pharmaceutical composition in the form of a solution and a second container comprising an aqueous liquid vehicle. At the time of administration, the contents of second container can be added to the contents of the first container or vice versa with mild agitation or shaking, resulting in the formation of nanodispersion.
While the present invention is disclosed generally above, additional aspects are further discussed and illustrated with reference to the examples below. However, the examples are presented merely to illustrate the invention and should not be considered as limitations thereto.

EXAMPLE 1
Table 1

Ingredients Comparative example 1 Comparative example 2 Example 1a Example lb

mg/g %w/w mg/g %w/w mg/g %w/w mg/g %w/w
Docetaxel 60 6 90 9 90 9 90 9
Caprylic acid 4.0 0.4 6.0 0.6 6.0 0.6 6.0 0.6
Sodium cholesteryl
Sulfate 4.0 0.4 6.0 0.6 6.0 0.6 6.0 0.6
Povidone (K-17) 50 5.0 75 7.5 75 7.5 75 7.5
Dehydrated alcohol 60 6.0 90 9.0 90 9.0 90 9.0
Paclitaxel ~ -- 2.5 0.25 5 0.5 10 1
Polyethylene glycol 400 q.s 1 gm q.s 100 q.s
1gm q.s 100 q.s 1 gm q.s 100 q.s 1 gm q.s 100
Appearance Clear colorless slightly viscous solution
Docetaxel, sodium cholesteryl sulfate, caprylic acid, povidone (K-17) and paclitaxel were weighed accurately in a glass vessel. Contents were dissolved in the required quantity of dehydrated alcohol and PEG-400 with stirring to obtain a clear concentrated drug solution. The solution was filtered through 0.2µ PVDF membrane filter. The pharmaceutical composition in the form of a solution so prepared was found to be clear colorless yellowish slightly viscous solution. The required amount of the pharmaceutical composition in the form of a solution was dispersed in the dextrose solution (5% w/v) with gentle shaking to get a translucent nanodispersion of drug having concentration of 0.5 mg/ml. Nanodispersion was analyzed for the tests such as: appearance, pH (Mettler Toledo-seven easy, pH Meter) and particle size (Nano-ZS, Malvern Particle size analyzer) described in table 2(a). The stability of the nanodispersion in terms of the particle size of the dispersed particles was determined initially as well as on storage.

Table 2: Observations on the aqueous dispersion obtained from the pharmaceutical composition in the form of a solution of example la, example lb at a concentration of 0.5 mg/ml in 5 % w/v dextrose solution

Example la Example 1b
Description Oh Almost clear to translucent dispersion Almost clear to dispersion translucent

8h Translucent dispersion with cloudy particles Almost dear to dispersion translucent
Particle size (mean in nm) Oh 82.0 68.9

2h 102.0 69.3

3h 188.0 ND

4h - 70.6

6h - ND

8h - 75.3
pH Oh - 4.00

8h - 4.00
Zeta potential (mV) - -27.7
Remarks Physically stable for 3 hours Physically stable for 8 h
It may be noted that when solution of docetaxel without any close structural analog, paclitaxel was dispersed in the aqueous vehicle for example, 5 % dextrose solution in purified water, although the dispersion showed particle size of less than 200 nm, the dispersion did not remain stable for more than 2 hours, But when solution of docetaxel with the close structural analog, paclitaxel, was dispersed in 5 % dextrose solution in purified water, the dispersion remained stable for about 3 hours. There was no aggregation or precipitation of docetaxel. When the amount of the close structural analog was further increased, the solution when dispersed in aqueous vehicle remained stable for extended period such as 8 hours. This effect of extended period of physical stability in comparison to the solution without the close structural analog was indeed surprising.

EXAMPLE 2
Table 3

Ingredients Comparative example 3 Examp Ie2

mg/g %w/w mg/g %w/w
Fenofibrate 130 13 130 13
Fenofibric acid - ~ 13 1.3
Caprylic Acid 6.0 0.6 6.0 0.6
Sodium Cholesteryl
Sulfate 6.0 0.6 6.0 0.6
Povidone (K-90) 75 7.5 75 7.5
Dehydrated alcohol 200 20 200 20
Polyethylene glycol 8000 200 20 200 20
DMSO 200 20 200 20
Polyethylene glycol 400 q.s 1 gm q.s 100 q.s 1 gm q.s 100
Appearance Hazy mixture Colorless slightly solution viscous clear
Fenofibrate, sodium cholesteryl sulfate, caprylic acid, povidone (K-90), polyethylene glycol 8000 and fenofibric acid were weighed accurately in a glass vessel. Contents were dissolved in the required quantity of dimethyl sulfoxide, dehydrated alcohol and PEG-400 with stirring to obtain a clear concentrated drug solution. The solution was filtered through 0.2µ PVDF membrane filter. The pharmaceutical composition in the form of a solution so prepared was found to be clear colorless yellowish slightly viscous solution. The required amount of the pharmaceutical composition in the form of a solution was dispersed in the water for injection with gentle shaking to get a translucent nanodispersion of drug having concentration of 1.3 mg/ml. Nanodispersion was analyzed for the tests such as: appearance, pH (Mettler Toledo-seven easy, pH Meter) and particle size (Nano-ZS, Malvern Particle size analyzer), described in table 4.

Table 4: Observations on the aqueous dispersion obtained from the pharmaceutical composition of example 2 at a concentration of 1.3 mg/ml in water for injection

Example 2
Description 0h Translucent dispersion

1h 30 min Settlement due to aggregates, change in appearance
Particle Size (mean in nm) Oh 382

lh 510
pH Oh 4.51
Zeta potential (mV) -36.4
Remarks Physically stable for 1hour
The stability of the nanodispersion in terms of the particle size of the dispersed particles was determined initially as well as on storage. The solution without fenofibric acid, a close structural analog did not provide a clear solution rather a hazy mixture was obtained. This indicates that the presence of structural analog of the fenofibrate, fenofibric acid, provides a clear solution. Further, when this solution is dispersed in an aqueous vehicle, a nanodispersion that is physically stable for a period of about 1 to 2 hours is obtained. The composition of water insoluble active ingredients like fenofibrate is meant for oral administration. The nanodispersion of such compositions are found to be physically stable for a period of about 1 to 2 hours, during which time a sufficient absorption of the drug through the mucosa is believed to take place.

EXAMPLE 3
Table 5

Ingredients Comparative Example 4 Comparative Example 5 Example 3
mg/g %w/w mg/g %w/w mg/g %w/w
SN-38 9 0.9 7.3 0.73 10 1
Caprylic acid 8.3 0.83 4.15 0.415 8.3 0.83
Sodium cholesteryl sulfate 6.7 0,67 3.35 0.335 6,7 0.67
Povidone (K-17) 75 7.5 75 7.5 75 7.5
Dehydrated alcohol 90 9 90 9 90 9
Dimethylsulfoxide 100 10 100 10 100 10
Irinotecan — - 1.75 0.175 5 0.5
Polyethylene glycol 400 q.s 1 gm q.s to 100 q.s 1 gm q.s to 100 q.s 1 gm q.s to 100
Appearance Yellowish slightly visco us solution
SN-38, sodium cholesteryl sulfate, caprylic acid, povidone (K-17) and irinotecan were weighed accurately in a glass vessel. Contents were dissolved in the required quantity of dimethyl sulphoxide, absolute alcohol and PEG-400 with stirring by heating the solution at 100°C to obtain a clear concentrated drug solution. The solution was filtered through 0.2u PVDF membrane filter. The pharmaceutical composition in the form of a solution so prepared was found to be clear colorless yellowish slightly viscous solution. The required amount of the pharmaceutical composition in the form of a solution was dispersed in the dextrose solution (5% w/v) with gentle shaking to get a translucent nanodispersion of drug having concentration of 0.1 mg/ml. Nanodispersion was analyzed for the tests such as: appearance, pH (Mettler Toledo-seven easy, pH Meter) and particle size (Nano-ZS, Malvern Particle size analyzer) described in table 6. The stability of the nanodispersion in terms of the particle size of the dispersed particles was determined initially as well as on storage i.e. after 48 h.

Table 6: Observations on the aqueous dispersion obtained from the preconcentrate of example 3 at a concentration of 0.1 mg/ml in 5% w/v dextrose solution

Time in hours Example 3
Description 0 Almost clear to translucent dispersion

48 Almost clear to translucent dispersion
Particle size (mean in nm) 0 258

1 266

4 266

6 256

8 260

48 273
pH 0 4.29

48 4.46
Zeta potential (mV) -5.35
Remarks Physically stable for 48 h.
It may be noted that the pharmaceutical composition in the form of a solution which is without a structural analog, irinotecan, when dispersed in the aqueous vehicle, was found to provide cloudy translucent dispersion. The dispersion was not suitable for administration particularly, the comparative example 4 showed presence of particles of about 953 nm whereas the comparative example 5 that included lesser than sufficient amount of irinotecan, showed aggregation in about 10 minutes, making it unsuitable for intravenous administration. On the contrary, the solution of SN-38 having sufficient amount of irinotecan, when dispersed in aqueous vehicle, the dispersion remained stable for about 48 hours i.e. there was no aggregation of particles. This effect of presence of sufficient amount of close structural analog, was indeed surprising and unexpected and also, very encouraging.

EXAMPLE 4
Table 7 (a)

Ingredients Quantity

mg/g (%w/w)
SN-38 7.3 0.73
Irinotecan 3.5 0.35
Caprylic acid 4.15 0.415
Cholesteryl sulphate 3.35 0.335
Povidone (K-17) 100 10
Dehydrated Alcohol 90 9
PEG-400 q.s 1 gm q.s to 100
Appearance Yellowish slightly viscous solution
Table 7(b): Observations on the aqueous dispersion obtained from the pharmaceutical composition in the form of a solution of example 4 at a concentration of 0.1 mg/ml in 5 % dextrose solution with 0.05 % dextran 40

Description Example 4

Almost white to pale yellow translucent dispersion
Particle size (mean in nm) Oh 305.2

2h 297.8

4h 300.0

6h 321.9

24 h 327.0
pH 4.44
Remark Physically stable for 24h
The pharmaceutical composition in the form of a solution of the present invention was prepared as described in example 1. The nanodispersion of pharmaceutical composition in the form of a solution produced was almost clear to slight yellow in appearance and had a mean particle size of 305.2nm.

EXAMPLE 5
Table 8 (a)

Ingredients Quantity

mg/g (%w/w)
SN-38 7.3 0.73
Irinotecan 3.5 0.35
Caprylic acid 4.15 0.415
Cholesteryl sulphate 3.35 0.335
Povidone (K-17) 100 10
Citric acid 0.282 0.0282
Dehydrated alcohol 90 9
PEG-400 q.s 1 gm q.s 100
Table 8(b): Observations on the aqueous dispersion obtained from the pharmaceutical composition in the form of a solution of example 5 at a concentration of 0.1 mg/ml in 5 % dextrose solution with 0.05 % dextran 40

Time in hrs Example 5
Description 0 Almost white to pale yellow translucent dispersion
Particle size (mean in nm) 0 372.9

2 385.8

4 384.0

24 376.8
pH 4.28
Remark Physically stable for 24h
The pharmaceutical composition in the form of a solution of the present invention was prepared as described in example 1. The nanodispersion of pharmaceutical composition in the form of a solution produced was almost clear to slight yellow in appearance and had a mean particle size of 372.9nm.

EXAMPLE 6
Table 9 (a)

Ingredients Example 6a Example
(Test) 6b Example (Placebo) 6c Example 6d (Control)

mg/g %w/w mg/g %w/w mg/g %w/w mg/g %w/w
SN-38 7.3 0.73 7.3 0.73 - - - -
Irinotecan 3.5 0.35 3.5 0.35 3.5 0.35 - -
Caprylic acid 8.30 0.83 4.15 0.415 4.15 0.415 4.15 0.415
Sodium cholesteryl sulphate 6.70 0.67 3.35 0.335 3.35 0.335 3.35 0.335
Povidone (K-17) 75 7.5 75 7.5 75 7.5 75 7.5
Dimethyl sulfoxide 100 10.0 100 10.0 100 10.0 100 10.0
Dehydrated alcohol 90 9.0 90 9.0 90 9.0 90 9.0
PEG-400 q.s 1
gm q.s 100 q.s 1 gm q.s 100 q.s 1 gm q.s 100 q.s 1 gm q.s 100
Appearance Clear solutions
Table 9 (b): Observations on the aqueous dispersion obtained from the pharmaceutical composition in the form of a solution of example 6a and Example 6b at a concentration of 0.1 mg/ml in 5 % dextrose solution with 0.05% dextran

Examples Example 6a Example 6b
Description Almost white to pale yellow translucent dispersion
Particle size (mean in nm) Oh 323.5 215.0

2h - 259.0

4h 330.9 254.0

6h - 297.7

8h 366 -

24 h 373.7 293.0
pH 4.4 4.28
Remarks Physically stable for 24ho urs
The pharmaceutical composition in the form of a solution of the present invention was prepared as described in example 1. The nanodispersion of pharmaceutical composition in the form of a solution produced was almost clear to slight yellow in appearance and had a mean particle size of 293 nm.

EXAMPLE 7
Pharmaceutical composition in the form of a solution of SN-38 prepared according to example 6b, example 6c (placebo) and example 6d (control) were prepared by a similar process as mentioned for example 3. These were subjected to efficacy studies using three xenograft tumor models using athymic nude Balb/c mice. The three human tumor cell lines that were utilized in these xenograft models were a) Human Mammary Carcinoma (MX-1), b) Colorectal Adeno Cell Carcinoma (HT-29) and c) Prostate Carcinoma (PC-3). The tumors of size 2 x 2mm were implanted subcutaneously in athymic nude Balb/c mice. For all these models samples were prepared and procedures described below are followed.
0.36 ml of pharmaceutical composition in the form of a solution was withdrawn using a 20 gauge needle attached to appropriately graduated syringe and to it was added 5.4 ml diluent vial (0.05% dextran 40 in 5% dextrose injection USP) to get a final concentration of 0.5 mg/ml of SN-38. The solution was shaken gently to obtain nanodispersion, which was used within half an hour of reconstitution. The placebo having irinotecan at a concentration of 0.24mg/ml and the control were also prepared similarly. The solutions to be administered were prepared keeping the dose volume of 8 ml/kg for pharmaceutical composition in the form of a solution of example 6b, placebo and control. Calculated dose of solution was given intravenously using 31 gauge needle attached to appropriately graduated syringe on days 0, 1, 2, 3 and 4. The injection was repeated in the same way for each animal in all other groups. Tumor measurement was also recorded on day 3.
Pharmaceutical composition in the form of a solution of example 6b following 5 injections once daily dosing in Human Mammary Carcinoma (MX-1), Human Colorectal Carcinoma (HT-29) and Human Prostate Carcinoma (PC-3) bearing athymic nude mice with Placebo (example 6c), control (example 6d) was studied as follow: a) Experimental procedure
Veterinary health check was performed to select the healthy animals at the time of receipt of animals. Selected animals were transplanted with Human Mammary Carcinoma (MX-1), Colorectal Adeno Cell Carcinoma (HT-29) and Prostate Carcinoma (PC-3) xenografts by procedure as described below.

1) Preparing tissue from donor animal for transplant
The entire experimental operations were carried out under Airstream Biological Safety Cabinet (Make: ESCO; Model: Airstream Class II E Series; Instrument ID: BRP/201) located in specific pathogen free barrier facilities. All surgical used for the transplantation procedure were autoclaved prior to use.
i) Media working stock was prepared by adding penicillin/streptomycin solution
(PEN/STREP; Lonza) in a 1:100 dilution to growth media. ii) The donor animal having a tumor diameter of around 10 mm was euthanized
with high dose of isoffurane (Isorane®) and the tumor was removed. iii) The tumor was then transferred to a sterile Petri dish containing media plus
penicillin/streptomycin. iv) The necrotic material, if any, from the tumor was removed and tumor fragments of around 2x2 mm were prepared
2) Transplant of tumor tissue
i) The recipient animal was anaesthetized using isofluane(Isorane®). ii) Using a cotton swab saturated with 70% (v/v) ethanol, the area from the mid-spine to the base of the tail was wiped. iii) A small, horizontal incision of around 5 mm in length, approximately 10 mm
above the base of the tail, was made using small surgical scissors. iv) The tip of the scissors was inserted into the incision, directly over the flank,
and the scissors opened slightly to introduce a pocket in the subcutaneous
space. v) One individual piece of tumor was inserted into the pocket created using
forceps. vi) The incision site was closed with one drop of 3M VetbondTM (Make: 3M
Animal Care Products) tissue adhesive. vii) After drying Neosporin® (Make: GlaxoSmithkline) antibiotic powder was
applied on the sealed incision. viii) Once the donor tumor was removed, the transplantation procedure was
completed within 30 minutes.

Tumor volume of the animals at the start of study was around 65-180 mm3. On day 0, tumor bearing animals were assigned randomly into 4 different groups of 10 animals. The tumor diameter (mm) was measured in two perpendicular diameters (Dl and D2) using digital Vernier Caliper (Make: Mitutoyo Digimatic Caliper; Model: CD-6"CSX; Instrument ID: BRP/099). The tumor volume was calculated using the formula for a sphere [(Dl+D2)/2]3 x0.5236 mm3. Each animal was weighed using a digital weighing balance. Tumor measurement was also recorded on day 3.
Tumor volume and body weight were then recorded twice weekly every week till the completion of study (from day 5 to day 42). The animals were checked daily throughout the study for deaths. The body weight, tumor volume and mortality data of treatment groups was compiled and analyzed using appropriate statistical method. For humane reasons animals were euthanized when implanted tumor volume reached >4,000 mm3. At completion of day 42 all animals were euthanized and handed to LAR. b) Evaluation of anti-tumor Activity
The efficacy of test and reference formulations was assessed and compared based on
following parameters,
Percentage T/C = (Mean Tumor volume of drug treated group on day X) x 100
(Mean Tumor volume of control group on day X)
The optimal value is the minimal T/C ratio that reflects the maximal tumor growth inhibition achieved. According to National Cancer Institute (NCI) criteria, T/C < 42% is considered for minimum acceptable anti-tumor activity, T/C < 20% is considered for moderate anti-tumor activity, T/C < 10% is considered to indicate highly significant anti-tumor activity.
Tumor regression was recorded as partial (PR) if the tumor volume decreased to less than 50% of the tumor volume at the start of the treatment without dropping below measurable size, or as complete (CR) if the tumor burden has become impalpable. Tumor regression in an animal experimental tumor models are an important end points in clinical relevance.

Specific tumor growth delay (SGD) is defined as being the difference in time for drug treated and control tumors to reach a given volume (V) divided by the time for the control tumors to reach the same volume (V),
SGDv = [Median days Tv (drug treated Eroup)-Median days Tv (control groups)]
Median days Tv control group Where V value being tumor volume after two volume doublings from initial tumor volume at the start of treatment and Tv being the time for drug treated or control groups to reach the given volumes. However, if V value was not achieved in test or reference group animal until last day of experiment, same value was considered as Tv for that animal.
The efficacy criteria for SGD parameter is >1, The mouse body weight changes = (Mouse wt. on day X - Mouse wt. on day 0) x 100
(Mouse wt. on day 0) A dose producing a weight loss > 15% of initial body weight is considered toxic. The recovery of weight after the completion of treatment will also be evaluated.
c) Survival analysis was done by Kaplan-Meier method. p<0.05 was considered as statistically significant.
d) Statistical analysis
All statistical analysis was carried with PRISM (GraphPad version 5.03, December 10, 2009) and p<0.05 was considered as statistically significant. Tumor volume and body weight data were analyzed by two-way and one-way ANOVA respectively. The Kaplan-Meier method was used to estimate survival and differences were analyzed by log-rank test.
Results of the in vivo tests are depicted graphically for Human Mammary Carcinoma (MX-1) in figure 1, Colorectal Adeno Cell Carcinoma (HT-29) in figure 2 and Prostate Carcinoma (PC-3) in figure 3. It may be concluded that that the pharmaceutical composition in the form of a solution of the present invention showed acceptable activity to highly significant anti-tumor activity in these models against the placebo treated groups.
It may be concluded that a highly significant anti-tumor activity was seen in the tested pharmaceutical composition in the form of a solution of the present invention at a dose

level of 4 mg/kg in Human Mammary Carcinoma (MX-1) xenograft in athymic nude mice. Also, an acceptable anti-tumor activity was seen in the tested pharmaceutical composition in the form of a solution of the present invention at a dose level of 4 mg/kg in Human Colorectal Carcinoma (HT-29) xenograft in athymic nude mice as well as an acceptable anti-tumor activity was seen in the tested pharmaceutical composition in the form of a solution of the present invention at a dose level of 4 mg/kg in Human Prostate Carcinoma (PC-3) xenograft in athymic nude mice.
EXAMPLE 8
Pharmaceutical composition in the form of a solution containing SN-38 (7-ethyl 10-hydroxy camptothecin) as described in example 6b was checked to determine the pharmacokinetics in Wistar rats. The pharmaceutical composition in the form of a solution of Example 6 (b) and placebo (example 6c) was prepared as mentioned in example 7. The sample preparation for the pharmacokinetic studies was prepared as follows: 9 ml of pharmaceutical composition in the form of a solution of example 6b was withdrawn using a 20 gauge needle attached to appropriately graduated syringe and added to 15 ml of diluent vial (0.05% dextran 40 in 5% dextrose injection USP) to get a final concentration of 3 mg/ml of SN-38. The solution was shaken gently to obtain nanodispersion, which was used within half an hour of reconstitution. The placebo (example 6c) sample preparation was done similarly.
Dose volume for test and placebo were 0.83 ml/kg, 1.67 ml/kg, 3.33 ml/kg for the dose 2.5mg/kg, 5 mg/kg and 10 mg/kg respectively. Test and Placebo were injected using 28.5 gauge needle attached to appropriately graduated syringe. a) Experimental procedure
Already received animals from LAR were used for the study. After acclimatization, on day 0, animals were weighed and the solution to be injected were prepared keeping the dose volume 0.83ml/kg, 1.67 ml/kg, 3.33 ml/kg at the dose of 2.5mg/kg and 10mg/kg respectively for placebo and test as 2.5 mg/kg, 5 mg/kg, 10 mg/kg. Single calculated dose of the drug solution was injected intravenously. The injection was repeated in the same way for each animal for other groups. Animals were anesthetized using Isoflurane (Compact Anesthesia Work Station, instrument ID BRP/164) and approximately 500 µL

of blood was collected by retro-orbital plexus puncture using heparinised capillaries in labeled (label includes Study number, Anima ID, Time point, Group) micro centrifuge tubes containing anticoagulant (15uL of 10% EDTA) at 0.083, 0.167, 0.333, 0.5, 1, 1.5, 2, 3, 6, 8 and 24 hour post intravenous injection. Plasma was separates from the collected blood samples by centrifugation at approximately 3000 rpm for 10 min at 10°C and was transferred into other labeled (label includes Study number, Anima ID, Time point, Group) micro centrifuge tubes containing 2% H3PO4 (PIasma:H3PO4 was 2:1 ratio). Plasma was stored at about -70°C (±10°C) in deep freezer, until analysis using available LC-MS/MS ADD. All the plasma samples were transferred in frozen condition in thermo cool box with ice packs to the test site. Plasma concentration data was provided by ADD for final report preparation.
b) Sample analysis
Plasma samples were stored at or below -70°C (±10°C) after receipt and analysed by available LC-MS/MS method. LOQ for SN-38 or irinotecan in plasma was 50ng/ml.
c) Pharmacokinetic analysis
Following parameters were determined from the plasma concentration obtained after
analysis using WinNonlin software version 5.0 by non-compartmental model.
AUCo-t: Area under concentration-time curve from time zero to time of last nonzero
concentration
The results of the study are tabulated as follows:
Table 10: Pharmacokinetic study of pharmaceutical composition in the form of a solution
of example 6b

Dose
(mg/kg) Cmax

(ng/ml) AUCJast (h*ng/ml) AUCinf (h*ng/ml)
2.5 1009.36 184.9 214.02
5 2163.15 954.33 1081.35
10 4499.08 2769.85 3143.37
The result shows a dose linearity of the pharmacokinetics parameters of SN-38 i.e. the exposure of SN-38 is linearly proportional to the dose administered.

We claim:
1. A pharmaceutical composition in the form of a solution comprising water insoluble therapeutically active ingredient and a close structural analog of the water insoluble active ingredient, one or more pharmaceutically acceptable excipients and one or more water miscible solvents wherein the water insoluble active ingredient is present in therapeutically effective amounts and the close structural analog is present in sufficient amounts such that when the composition is added to an aqueous vehicle, it forms a nanodispersion suitable for intravenous infusion wherein said nanodispersion is stable for longer periods as compared to a dispersion formed from an identical composition but devoid of the close structural analog of the water insoluble active ingredient.
2. A pharmaceutical composition according to claim 1, wherein pharmaceutically acceptable excipient is selected from the group consisting of water soluble polymers, surfactants and mixtures thereof.
3. A pharmaceutical composition as claimed in claim 1, wherein the water miscible solvent is selected from the group consisting of alcohols, glycols, dimethyl sulfoxide and mixtures thereof.
4. A pharmaceutical composition according to claim 1, wherein water insoluble therapeutically active ingredient is SN-38 and the close structural analog is irinotecan.
5. A pharmaceutical composition according to claim 4, wherein the aqueous vehicle contains one or more colloid stabilizers.
6. A pharmaceutical composition according to claim 1 wherein water insoluble therapeutically active ingredient is docetaxel and the close structural analog is paclitaxel.