FORM 2
THE PATENTS ACT, 1970
(39 OF 1970)
PROVISIONAL SPECIFICATION
(See section 10)
'SUSTAINED RELEASE PHARMACEUTICAL COMPOSITIONS'


SUN PHARMACEUTICAL INDUSTRIES LTD.

A company incorporated under the laws of India having their office at ACME PLAZA, ANDHERI-KURLA ROAD, ANDHERI (E), MUMBAI-400059, MAHARASHTRA, INDIA
The following specification describes the nature of this invention.


SUSTAINED RELEASE PHARMACEUTICAL COMPOSITION
The present invention relates to sustained release pharmaceutical microsphere compositions comprising goserelin acetate.
The present invention provides a sustained release microsphere composition comprising microspheres comprising (A) a biodegradable polymer which is a copolymer of lactic acid and glycolic acid having a monomer ratio in the range of about 1:1 to about 3:1, and (B) a therapeutically effective amount of goserelin acetate, and pharmaceutically acceptable excipients. The biodegradable polymer preferably has an average molecular weight within the range of about 10,000 to about 200,000. The microspheres of the present invention have a volume mean diameter in the range of about 10 microns to about 20 microns.
Heretofore, goserelin has been administered through subcutaneous injection of implants containing goserelin, whereby the goserelin is slowly released in vivo over one month or three months from the implants. The commercially available goserelin implants are not suitable for intramuscular administration. We have found goserelin microspheres of the present invention to be useful in that they can be administered by intramuscular injection and provide a slow release of goserelin over a period of about one month or about three months.
The present invention may be summarized as follows -
(a) A sustained release microsphere composition comprising -
(i) microspheres comprising (A) a biodegradable polymer which is a copolymer of lactic acid and glycolic acid having a monomer ratio in the range of about 1:1 to about 3:1, and (B) a therapeutically effective amount of goserelin acetate, and
(ii) pharmaceutically acceptable excipients.
(b) A sustained release microsphere composition as described in (a) above, wherein the biodegradable polymer used has an average molecular weight within the range of about 10,000 to about 20,000.
(c) A sustained release microsphere composition as described in (a) above, wherein the microspheres have a volume mean diameter in the range of about 10 microns to about 20 microns.

(d) A sustained release microsphere composition as described in (a) above, wherein the composition is capable of delivering goserelin acetate for a period of about one month or about three months.
(e) A sustained release microsphere composition comprising goserelin acetate, wherein the microsphere is suitable for intramuscular injection.
(f) A sustained release microsphere composition as described in (e) wherein the composition is capable of delivering goserelin acetate for a period of about one month or about three months.
(g) A sustained release microsphere composition as described in (a) above, wherein the pharmaceutically acceptable excipient is mannitol.
The examples that follow do not limit the scope of the present invention and are merely used as illustrations.
Example 1
A sustained release injection composition of goserelin acetate was obtained as described in Table 1 below. The goserelin microspheres of the present invention are obtained using a novel process.
Table 1

Ingredients Quantity (mg/vial)
Microsphere formulation
Goserelin acetate 3.6
Purified gelatin 1.3
DL-lactic acid and glycolic acid copolymer (3:1, molecular weight 10,000) 66.2
Mannitol 13.2
Formulation medium
Sodium carboxymethyl cellulose 5.0
Mannitol 50.0
Polysorbate 80 1.0
Water for injection q.s.
Glacial acetic acid q.s. for pH adjustment
Goserelin acetate was mixed with purified gelatin and the mixture was dissolved in water. The solution thus obtained was subjected to filtration, followed by lyophilisation of the solution to obtain a cake. This cake was dissolved in a sufficient amount of water for injection to obtain an aqueous phase. This aqueous phase was emulsified using a solution of the lactic acid-glycolic acid copolymer in methylene chloride, in a first tank, to obtain a primary emulsion. The primary emulsion was cooled to about 15°C for about 30 minutes, and then pumped to a second tank

containing an aqueous solution of mannitol and 0.1% polyvinyl alcohol. The mixture was homogenized to obtain a water/oil/water emulsion. The excess solvent was evaporated from this ternary emulsion, followed by sieving and drying of the microspheres. The dry microspheres are suspended in aqueous mannitol solution and lyophilized. The lyophilized microspheres are then filled into vials.
The lyophilized microspheres are then suspended in a formulation medium prior to administration, the medium comprising sodium carboxymethyl cellulose, mannitol and polysorbate 80 in sterile water for injection, the pH of the medium being adjusted with glacial acetic acid to about pH 5.0-6.0.
Example 2 A sustained release injection composition of goserelin acetate was obtained as described in Table 2 below.
Table 2

Ingredients Quantity (mg/vial)
Microsphere formulation
Goserelin acetate 3.6
Purified gelatin 1.3
DL-lactic acid and glycolic acid copolymer (1:1, molecular weight 20,000) 66.2
Mannitol 13.2
Formulation medium
Sodium carboxymethyl cellulose 5.0
Mannitol 50.0
Polysorbate 80 1.0
Water for injection q.s.
Glacial acetic acid q.s. for pH adjustment
The goserelin microspheres were obtained by a process similar to that mentioned in Example 1 above.
Example 3
The effect of varying ratios of lactic acid to glycolic acid in the polylactic-glycolic acid biodegradable polymers used to obtain the microspheres of the present invention was also studied. Two sustained release goserelin microsphere compositions were prepared by the process detailed in example 1 above, using (1) polylactic-glycolic acid copolymer having a lactic acid to glycolic acid ratio of 3:1, and (2) polylactic-glycolic acid copolymer having a lactic acid to glycolic acid ratio of 1:1.The physicochemical properties of the two compositions were found to

be comparable as regards the particle size, with both compositions comprising microspheres with volume mean diameter of about 15 microns. The in vitro release profile from these two compositions is recorded in Table 3 below.
Table 3

Time (days) % drug released from microspheres
obtained using polylactic-glycolic acid
polymer having lactic acid to glycolic
acid ratio of 3:1 % drug released from microspheres
obtained using polylactic-glycolic acid
polymer having lactic acid to glycolic
acid ratio of 1:1
1 28.47 18.78
14 59.88 47.68
21 75.78 87.09
28 83.35 99.09
Example 4
The effect of molecular weight of the polymer used to obtain the microspheres on the in vitro release of goserelin is summarized below. Two sustained release goserelin microsphere compositions were prepared by the process detailed in example 1 above, using polylactic-glycolic acid copolymer having a lactic acid to glycolic acid ratio of 3:1, and having a molecular weight of (1) 15,000 Daltons and (2) 10,000 Daltons. The in vitro release of goserelin from these two compositions is summarized in Table 8 below.
Table 4

Time (days) % drug released from
microspheres obtained using
polylactic-glycolic acid polymer of
molecular weight 15,000 Daltons % drug released from microspheres
obtained using polylactic-glycolic
acid polymer of molecular weight
10,000 Daltons
1 27.84 17.07
14 55.48 65.20
28 56.96 87.97
35 59.72 98.04
The microspheres obtained using the polymer with molecular weight of 15,000 were found to provide an initial burst of goserelin acetate, followed by a slow release, with only about 60% of the goserelin acetate released in 35 days. The microspheres obtained using the polymer with the lower molecular weight of 10,000 were found to provide a satisfactory release profile with about 98% of the goserelin acetate being released in 35 days. Thus, the polymer with lower molecular weight was found to be desirable.

Example 5
The goserelin acetate microsphere compositions of Example 1 and Example 2 were subjected to in vivo studies in rats. The study was done on male Wistar rats of weight 150 - 200gm. The rats were assigned randomly into groups having ten animals each, depending upon the number of samples to be tested.
On day 0, about 0.3 ml of blood sample was collected with rat haematocrit capillaries in 1.5ml eppendorfs apparatus from retro-orbital plexus from all the rats and centrifuged at 3000rpm for 15 minutes at 10°C. The supernatant was removed and transferred to another eppendorfs apparatus and recentrifuged again at 3000rpm for 15 minutes at 10°C. The serum samples thus obtained were assayed for levels of testosterone by ELISA.
On day 1, the animals were weighed and the dose volume calculated according to body weight. The test Goserelin acetate depot formulation was suspended in the diluent provided such that the final dose volume was lml/kg body weight. 0.1ml of suspension/lOOgm body weight was injected intramuscularly (i.m.)/subcutaneously (s.c.) in the animals. For vehicle control group, diluent was injected i.m./s.c. at the dose volume of lml/kg.
24 hours after injection (day 1 after injection), 0.3 ml of blood sample was collected with rat haematocrit capillaries from retro-orbital plexus of each animal in 1.5 ml eppendorfs apparatus. The blood samples were centrifuged at 3000rpm for 15 minutes at 10°C. The supernatant was removed & transferred to another eppendorfs and recentrifuged again at 3000rpm for 15 minutes at 10°C. The serum samples were assayed by ELISA for Testosterone levels.
The same procedure was repeated for collection of serum and analysis for testosterone levels by ELISA on days 3, 7, 14, 21, 28, 35 post injection for 1-month depot formulation and 3, 7, 14, 21. 28, 56, 84 and 112 days of post injection for 3-month depot formulation. On the last day of the study, the animals were weighed and sacrificed in carbon dioxide atmosphere and the prostate, testes (both) and seminals were collected aseptically and their wet-weight recorded.
The Testosterone values (ng/ml) and organ weight/body weight ratio of different groups was compared with that of vehicle control group. The testosterone concentration in the experimental animals as measured is recorded in Table 5 below.

Table 5

Time (days) Testosterone concentration (ng/ml)
in rats treated with goserelin
acetate microsphere composition of
Example 1 Testosterone concentration (ng/ml)
in rats treated with goserelin
acetate microsphere composition of
Example 2
0 1.47 4.76
1 3.5 8.96
3 0.81 0.44
7 0.02 0.04
14 0.08 0.14
21 0.03 0.21
28 1.51 0.19
35 0.34 2.37
Dated this 1st day of March, 2005.
MSWATI VEERA, PATENT ATTORNEY (IN-HOUSE), SUN PHARMACEUTICAL INDUSTRIES LIMITED.