FORM 2
THE PATENTS ACT, 1970 (39 of 1970)
PROVISIONAL SPECIFICTAION / COMPLETE
SPECIFICATION
(See section 10 and rule 13)
1. TITLE OF THE INVENTION
VACCINE ADJUVANTS

2. APPLICANT(S)
(a) NAME : CADILA PHARMACEUTICALS LTD
(b) NATIONALITY: An INDIAN Company
(c) ADDRESS : "Cadila Corporate Campus", Sarkhej - Dholka Road, Bhat, Ahmedabad -
382210, Gujarat, India
3. PREAMBLE TO THE DESCRIPTION
PROVISIONAL SPECIFICATION
The following specification describes invention.

COMPLETE SPECIFICATION
Tho following opocificotion particularly doocriboo tho invention ond tho manner in which it io to bo performed

4. DESCRIPTION
(Description starts from next page)


FIELD OF INVENTION
The invention relates to an adjuvant that can stimulate the mammalian immune system against a wide variety of antigen(s). The invention also provides novel adjuvants to potentiate effect of other adjuvants, their compositions and a novel method of identifying compounds for use as adjuvant(s).
BACKGROUND INFORMATION:
Adjuvants are used to
• Increase antibody response in magnitude or function
• Cell mediated immune response
• Reduction in antigen dose/time of mounting response
• Induction of mucosal immunity
The only adjuvants which are universally approved for human use are aluminum salts. Some of the known adjuvants progressing in clinic are
• Oil-in-water emulsions - MF59
• Saponins: QS21, ISCOMS
• CpG (Oligonucleotides) - TLR agonists
• Other TLR agonists and Innate Immunity
• Immunopeptides
The above mentioned adjuvants are at various stages of development. There is still a need to have novel adjuvants to provide novel vaccine with reduced antigen, reduced frequency, increased immunogenic potential. Currently available adjuvants are outcome of painstaking research in absence of a method to identify potential compound with adjuvant properties. OBJECT OF INVENTION:
The main object of invention is to provide adjuvant that can stimulate the mammalian immune system against a wide variety of antigen (s).
Another object of invention is to provide a novel method of identifying compounds for use as adjuvant(s).
Yet another object of invention is to provide novel adjuvants to potentiate effect of other adjuvants.
Yet another object of invention is to provide composition consisting novel adjuvants as per this invention for administration to mammals. DESCRIPTION OF DRAWING:
Figure: 01 Antibody titers following immunization with Hepatitus - B antigen with different dosage of BB as an adjuvant.
2

DETAILED DESCRIPTION:
Surprisingly compound 3pHydroxy-5,16-pregnadien.20-one (C2,H30O2) here in after called as BB is found to have adjuvant properties. When wistar rats are administered 10 ug of hepatitis-b surface antigen with or without BB in various concentrations, the antibody titer raised were higher when BB is combined with hepatitis antigen (Fig). On day 42 the effect is maximal and also dose dependent, increasing with increasing amount of BB.
3pHydroxy-5,16-pregnadien-20-one (BB) is a compound which is known FXR inhibitor. The structure of 3pHydroxy-5,16-pregnadien-20-one (BB) is as under: BB is water insoluble.

3β-Hydroxy-5,16-pregnadien-20-one The FXR inhibitors include following guggulipd, guggulsterones 3β-Acetoxypregna-5,16-dien-20-one (16-DPA), 4,16-Dienpregna-3,20-dione, 3p-Actoxypregna-5-en-20-one, 3p-Hydroxypregna-5-en-20-one, 5,16-Dien-pregnane-3,20-diol, 5,17(20)-Dien-pregna-3,16-diol-diacetate, 5,17(20)-Dien-pregna-3,16-diol, 3p-Hydroxypregna-5,16-dien-20-one, 7-{2-Hexyloxy-3,5-diisopropyl-phenyl)-3-methyl-octa"2,4,6-trienoit acid, Z-Guggulsterone, [4f17(20)-Cis-pregnadiene-3f16-dione], E-Guggulsterone [4,17(20-Cis-pregnadiene-3,16-dione] etc. The structures of these compounds are as under:




AcO

3p-Acetoxypregna- 5,16-dien-20-one (16-DPA)

3p-Hydroxypregna- 5,16-dien-20-one

4,16-Dienpregna-3,20-dione

3

3β-Actoxypregna-5-en-20-one

3β-Hydroxypregna-5-en-20- 5,16-Dien-pregnane-3,20-
diol

AcO

5,17(20)-Dien-pregna-3,16- 5,17(20)-Dien-pregna-3:16-
diol-diacetate diol

3β-Hydroxypregna- 5,16-dien-20-one

7-(2-Hexyloxy-3,5-diisopropyl- Z-Guggulsterone E-Guggulsterone
phenyl)-3-methyl-octa-2,4,6- 4.17(20)-Cis-pregnadiene-3,16- 4,17(20)-Cis-pregnadiene-3,l6-
trienoic acid dione dione
Some of them are known to have hypolipaemic effects. However they are not known to have adjuvant properties.
According to present invention vaccines adjuvant can be identified based FXR inhibition properties. BB potentiate effect of other adjuvants like Mycobacterium w, alum etc.
The pharmaceutical composition made using present invention can be a suspension, containing antigen In appropriate vehicle with BB suspended into it. It can be with or without preservatives.
Following are examples of compositions as per present invention without
preservatives
A. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen

4

3(3Hydroxy-5, 16-pregnadien-20-one (C21H30O2) 5 ugM
Water for injection I. P. q. s. to 1.0 ml
B. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10 μg
3pHydroxy-5,16-pregnadien-20-one (C21H30O2) 5 μgM
Normal Saline q. s. to 1.0 ml
C. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10 μg
3β Hydroxy-5,16-pregnadien-20-one(C21H3o02) 10 μgM
Water for injection ). P. q. s. to 1.0 ml
D. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10 μg
3pHydroxy-5, 16-pregnadien-20-one (C21H30 10 μgM
Normal Saline q. s. to 1.0 ml
E. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10 μg
3pHydroxy-5,16-pregnadien-20-one (C21H30O2) 20 μgM
Water for injection I. P. q. s. to 1.0 ml
F. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10 μg
3pHydroxy-5,16-pregnadien-20-one {C21H30O2) 20 μgM
Normal Saline q. s. to 1.0 ml
G. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10μg
3pHydroxy-5, 16-pregnadien-20-one (C21H30O2) 50 μgM
Water for injection I. P. q. s. to 1.0 ml
H. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 10 μg
3pHydroxy-5, 16-pregnadien-20-one (C2iH30O2) 50 μgM
Normal Saline q. s. to 1.0 ml
I. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 20 μg
3pHydroxy-5, 16-pregnadien-20-one (C21H30O2) 5 μgM
Water for injection I. P. q. s. to 1.0 ml
J. Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 20 μg
5

3β Hydroxy-5,16-pregnadien-20-one (C21H30O2) Normal Saline
K.
Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 3β Hydroxy-5,16-pregnadien-20-one (C21H30O2) Water for injection i. P.
L.
Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 3β Hydroxy-5, 16-pregnadien-20-one (C21H30O2) Normal Saline
M.
Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 3β Hydroxy-5,16-pregnadien-20-one (C2iH3o02) Water for injection I. P.
N.
Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 3β Hydroxy-5, 16-pregnadien-20-one (C21H30O2) Normal Saline
O.
Each 1.0 ml of composition contains:
Recombinant Hepatitis B surface antigen 3β Hydroxy-5,16-pregnadien-20-one (C21H30O2) Water for injection I, P.
P.
Each 1.0 mi of composition contains:
Recombinant Hepatitis B surface antigen 3β Hydroxy-5; 16-pregnadien-20-one (C21H30O2) Normal Saline

5 μgM
q-s to 1.0 ml
20 μg
10μgM
q-s. to 1.0 ml
20 μg
20μgM
q-s, to 1.0 ml
20 μg
20μgM
q- s. to 1.0 ml
20μg
50μgM
9-s to 1.0 ml)
20 μg 50 μgfyi
qs. to 1.0 ml
20μg 50 μgly|
q. s. to 1.0 ml

Date: 26-Sep-08

6


Date: 26-Sep-08

FIGURE 1
Sheet 01 of 01
For Cadila Pharmaceuticals Ltd.,

Dr. Bakules'h M. Khamar Executive Director - Research

2 9 SEP 2008