Title of Invention	AN EXOGENOUS PROTEIN-FREE PULMONARY SURFACTANT COMPOSITION FOR ADULT RESPIRATORY DISTRESS SYNDROME AND A PROCESS FOR PREPAIRING THE SAME
Abstract	This invention relates to an exogenous protein-free surfactant composition for use in treating adult respiratory distress syndrome (ARDS) a process for preparing the surfactant composition. The composition contains alteast one phospholipid sufactant and a natural oil selected from eucalyptus oil and its derivatives. This composition prevents the interaction between surfactants and hematological agents and will thus be effective in ARDS. It is also cost effective and does not have tha side-effects normally seen when protein containing surfactants are used.

Title of Invention

AN EXOGENOUS PROTEIN-FREE PULMONARY SURFACTANT COMPOSITION FOR ADULT RESPIRATORY DISTRESS SYNDROME AND A PROCESS FOR PREPAIRING THE SAME

Abstract

This invention relates to an exogenous protein-free surfactant composition for use in treating adult respiratory distress syndrome (ARDS) a process for preparing the surfactant composition. The composition contains alteast one phospholipid sufactant and a natural oil selected from eucalyptus oil and its derivatives. This composition prevents the interaction between surfactants and hematological agents and will thus be effective in ARDS. It is also cost effective and does not have tha side-effects normally seen when protein containing surfactants are used.

Full Text	FORM 2 THE PATENTS ACT 1970 (39 of 1970) & THE PATENTS RULES, 2003 COMPLETE SPECIFICATION (See Section 10; rule 13) TITLE OF THE INVENTION An exogenous protein-free pulmonary surfactant composition for Adult Respiratory Distress Syndrome and a process for preparing the same APPLICANT Indian Institute of Technology, Bombay, Powai, Mumbai 400 076, Maharashtra, India, an autonomous educational institute established in India under the Institutes of Technology Act 1961 INVENTORS Under Section 28(2) Dr. Rinti Banerjee , Ms Rachana, Dr. Jayesh Bellare, Dr Ram Puniyani all Indian nationals of Indian Institute of Technology, Bombay, Powai, Mumbai 400 076, Maharashtra, India The following complete specification particularly describes the nature of the invention and the manner in which it is to be performed. FIELD OF INVENTION This invention relates to an exogenous protein-free pulmonary surfactant composition for Adult Respiratory Distress Syndrome (ARDS) and a process for preparing the same. BACKGROUND OF INVENTION Endogenous pulmonary surfactant which is normally present and active in the system enables gas exchange during respiration. This natural surfactant is a complex mixture of many components such as dipalmitoyl phosphatidyl choline, lecithins, phosphatidyl glycerol, cholesterol, surfactant specific proteins, phosphatidyl ethanolamine, glycerides, phosphatidyl serine, lysolecithin and fatty acids. In ARDS, several hematological (derived from blood) agents like plasma proteins and red cell membranes flood the alveoli due to capillary damage and these prevent the lung surfactant from functioning. Due to inactivation of natural pulmonary surfactants, surface tension at the air-alveolar interface in the lungs increases, resulting in inadequate gas exchange during respiration. In order to permit adequate gas exchange in the lungs, rate of breathing and respiratory pressures are increased to re-expand the alveoli with each intake of breath. This condition is life threatening. 2 This condition differs from the Neonatal Respiratory Distress Syndrome (NRDS) that occurs in premature infants and which is caused by a deficiency of lung surfactant. Thus, the surfactant therapy in NRDS may not be effective for ARDS as this is not a simple deficiency state. Surfactant replacement therapy has had only partial benefits in treating ARDS and other surfactant inhibitory states. Most studies have shown no beneficial effects or non-significant benefits. Exogenous surfactants used as replacement in NRDS are rarely effective in ARDS. This is because the surfactants are unable to prevent surfactant interactions with hematological agents like plasma proteins and "red cell membranes. Further, many of these formulations are animal derived surfactants obtained mainly from cows, pigs and goats, which contain animal derived proteins. Animal derived synthetic surfactant compositions are more efficient in overcoming the inhibition of surfactant in ARDS than synthetic protein free products. However, proteins associated with these surfactants are immunogenic and may react with circulating anti-bodies in the system. Use of such surfactants have also shown incidence of increased intra ventricular hemorrhage. Moreover, animal derived surfactants are very expensive. As such these animal derived surfactants find little clinical use in ARDS. 3 Exogenous artificial surfactants are also known. Such compositions contain dipalmitoyl phosphatidyl choline hereinafter referred as (PC) in combination with phosphatidyl glycerol reference as (PG) in the ratio of 7:3. This composition may be reconstituted with sterile sodium chloride or Ringer's lactate solution. Compositions having PC in combination with co-surfactants such as tyloxapol and hexadecanol are also known in the art. These compositions are protein free However, these known protein free compositions exhibit very poor efficacy and cannot prevent the interaction of surfactants and hematological agents, cannot overcome the inhibition of surfactants in ARDS and are hence ineffective in ARDS. ARDS patients in India are not given surfactant therapy on a regular basis due to the non-availability of effective low cost surfactant replacements. Hence, the need to develop a surfactant replacement which avoids the drawbacks of animal derived surfactant and the need for an effective synthetic surfactant replacement composition for ARDS has been felt for sometime. OBJECT OF INVENTION The object of this invention is to provide an exogenous protein free pulmonary surfactant composition, which is effective even in the presence of inhibitory 4 agents like plasma proteins and red cell membranes and which does not exhibit any harmful side effects and is also cost effective. DESCRIPTION OF INVENTION and PREFRRED EMBODIMENTS This invention relates to an exogenous protein free pulmonary surfactant composition for treating adult respiratory syndrome which comprises atleast one phospholipid surfactant and a natural oil selected from eucalyptus oil or its constituents. This invention also includes a process for producing an exogenous protein free pulmonary surfactant composition for treating adult respiratory syndrome which comprises the steps of preparing a solution of atleast one phospholipid surfactant in an organic solvent, drying the said solution under vacuum, hydrating the residue thus obtained with aqueous isotonic electrolyte solutions and adding natural oils selected from eucalyptus oil or its constituents thereto. Phospholipid surfactant capable of being used in this composition is dipalmitoyl phosphatidyl choline (PC). The pH of the solution may be adjusted to 7 to 7.4 and the preferred organic solvent is chloroform : methanol (2:1). 5 Preferred composition may have 0.1 to 1% w/v of eucalyptus oil or its constituents. Phospholipid natural oil ratio may be 70:30 to 10:90 w/v lipids with final concentrations in the range of 0.1 to 1%. Eucalyptus oil used in the range of 5 to 6 microlites. Though eucalyptus oil is known to reduce phlegm in traditional medicines, it has not been used as a surfactant. None of the protein free surfactants can prevent interactions of the surfactants with the hematological agents. The uniqueness of this composition is that it contains one or two phospholipid constituents that are present in natural living surfactants which in the presence of eucalyptus oil or its constituents, prevents the interaction between surfactant and the hematological agents and hence overcomes inhibition achieving a function which is as good as natural surfactants in the absence of inhibitors. Efficacy of this composition has been tested and is found to be better than the hitherto known artificial surfactants. Surfactants compositions of this invention containing eucalyptus oil are as effective in the presence of hematological inhibitors as animal protein based surfactants are in their absence and are hence 6 of benefit. Further, they are without the harmful side effects of animal proteins. Cost factor is also an important aspect of this invention. Parameters for a surfactant to be effective for ARDS are (1) preventing interaction between surfactant and hematological agents (2) achievement of low minimum surface tension in the presence of hematological agents (3) low compressibility in the presence of hematological agents (4) requirement of a minimal area change for achievement of low surface tension in the presence of hematological agents. The composition of the subject invention fulfills all these conditions. The following examples are only illustrative and are not intended to limit the scope of this invention. Example 1: A solution of dipalmitoyl phosphatidyl choline in chloroform: methanol (2:1) in the concentration of 10 mg / ml was allowed to evaporate. To the residue 1 mL of normal saline was added and vigorously shaken. Eucalyptus oil 6 microl was then added to this solution. The solution was left undisturbed to allow hydration for a short period example 1 hr. The final phospholipids oil concentration in saline was 0.1%. 7 Example 2: The procedure of example 1 was followed using 12 hour hydration instead of 1 hour. Evaporation is performed to get a uniform thin film of PLS which facilitates uniform distribution of PLS in normal saline. Solvent in which PLS is to be dissolved may be chloroform, a mixture of chloroform and methanol in the ratio of 2:1. Other organic solutions may also be used in combination with methanol and / or chloroform. The hydration period controls the particle sizes of the formulations. Short periods of hydration for example 1 hour results in nanoparticles of 500-800 nm size which can be used in an inhalable form whereas longer periods of hydration for example 12 hours results in particles of a few microns which can be used as injections into the airways (intratracheal instillation). Any isotonic polyelectrolyte solution may be used for hydration. The polyelectrolytes should not contain calcium to prevent adverse interactions with hematological agents in ARDS. This invention described hereinabove and claimed in the appended claims do not exclude obvious equivalents, modifications and alternations. 8 WE CLAIM :- 1. An exogenous protein-free pulmonary surfactant composition for treating Adult Respiratory Syndrome comprising atleast one phospholipid surfactants and a natural oil selected from eucalyptus oil and/or its constituents. 2. The composition as claimed in claim 1 wherein the said composition is in the form of nanoparticles. 3. The composition as claimed in claim 1 wherein normal saline is used as a medium. 4. The composition as claimed in claims 1 to 3 wherein said phospholipid surfactant is dipalmitoyl phospahtidyl choline (PC). 5. The composition as claimed in claims 1 to 4 which contains 0.1 to 1% w/v of eucalyptus oil. 6. The composition as claimed in claims 1 to 5 wherein surface tension of composition on film compression in the presence of hematological agent is 0-5 mN/m. 7. A process for producing an exogenous protein-free pulmonary surfactant composition for treating adult respiratory syndrome comprising the steps of preparing a solution of atleast one phospholipid surfactants in an organic solvent, drying the same under vacuum, hydrating the residue obtained thereby with aqueous isotonic electrolyte solution and adding a natural oil selected from eucalyptus oil or its constituents thereto 9 The process as claimed in claim 7 wherein prior to hydration, the pH of the solution is adjusted to 7.2 to 7.4 The process as claimed in claims 7-8 wherein said composition has a phospholipid oil ratio of 70:30 to 10:90 w/v. An exogenous protein free pulmonary surfactant composition for treating adult respiratory syndrome substantially as herein described. A process for preparing exogenous protein-free surfactant composition for treating adult respiratory syndrome substantially as herein described Dated this 30th day of September 2005 10 ABSTRACT This invention relates to an exogenous protein-free surfactant composition for use in treating adult respiratory distress syndrome (ARDS) a process for preparing the surfactant composition. The composition contains alteast one phospholipid surfactant and a natural oil selected from eucalyptus oil and its derivatives. This composition prevents the interaction between surfactants and hematological agents and will thus be effective in ARDS. It is also cost effective and does not have the side -effects normally seen when protein containing surfactants are used.

Full Text

FORM 2
THE PATENTS ACT 1970
(39 of 1970)
&
THE PATENTS RULES, 2003
COMPLETE SPECIFICATION
(See Section 10; rule 13)
TITLE OF THE INVENTION
An exogenous protein-free pulmonary surfactant composition
for Adult Respiratory Distress Syndrome
and a process for preparing the same
APPLICANT
Indian Institute of Technology, Bombay, Powai, Mumbai 400 076, Maharashtra, India, an autonomous educational institute established in India under the Institutes of Technology Act 1961
INVENTORS
Under Section 28(2)
Dr. Rinti Banerjee , Ms Rachana, Dr. Jayesh Bellare, Dr Ram Puniyani all Indian nationals of Indian Institute of Technology, Bombay, Powai, Mumbai 400 076, Maharashtra, India
The following complete specification particularly describes the nature of the invention and the manner in which it is to be performed.

FIELD OF INVENTION
This invention relates to an exogenous protein-free pulmonary surfactant composition for Adult Respiratory Distress Syndrome (ARDS) and a process for preparing the same.
BACKGROUND OF INVENTION
Endogenous pulmonary surfactant which is normally present and active in the system enables gas exchange during respiration. This natural surfactant is a complex mixture of many components such as dipalmitoyl phosphatidyl choline, lecithins, phosphatidyl glycerol, cholesterol, surfactant specific proteins, phosphatidyl ethanolamine, glycerides, phosphatidyl serine, lysolecithin and fatty acids. In ARDS, several hematological (derived from blood) agents like plasma proteins and red cell membranes flood the alveoli due to capillary damage and these prevent the lung surfactant from functioning. Due to inactivation of natural pulmonary surfactants, surface tension at the air-alveolar interface in the lungs increases, resulting in inadequate gas exchange during respiration. In order to permit adequate gas exchange in the lungs, rate of breathing and respiratory pressures are increased to re-expand the alveoli with each intake of breath. This condition is life threatening.
2

This condition differs from the Neonatal Respiratory Distress Syndrome (NRDS) that occurs in premature infants and which is caused by a deficiency of lung surfactant. Thus, the surfactant therapy in NRDS may not be effective for ARDS as this is not a simple deficiency state.
Surfactant replacement therapy has had only partial benefits in treating ARDS and other surfactant inhibitory states. Most studies have shown no beneficial effects or non-significant benefits. Exogenous surfactants used as replacement in NRDS are rarely effective in ARDS. This is because the surfactants are unable to prevent surfactant interactions with hematological agents like plasma proteins and "red cell membranes. Further, many of these formulations are animal derived surfactants obtained mainly from cows, pigs and goats, which contain animal derived proteins. Animal derived synthetic surfactant compositions are more efficient in overcoming the inhibition of surfactant in ARDS than synthetic protein free products. However, proteins associated with these surfactants are immunogenic and may react with circulating anti-bodies in the system. Use of such surfactants have also shown incidence of increased intra ventricular hemorrhage. Moreover, animal derived surfactants are very expensive. As such these animal derived surfactants find little clinical use in ARDS.
3

Exogenous artificial surfactants are also known. Such compositions contain dipalmitoyl phosphatidyl choline hereinafter referred as (PC) in combination with phosphatidyl glycerol reference as (PG) in the ratio of 7:3. This composition may be reconstituted with sterile sodium chloride or Ringer's lactate solution. Compositions having PC in combination with co-surfactants such as tyloxapol and hexadecanol are also known in the art. These compositions are protein free However, these known protein free compositions exhibit very poor efficacy and cannot prevent the interaction of surfactants and hematological agents, cannot overcome the inhibition of surfactants in ARDS and are hence ineffective in ARDS.
ARDS patients in India are not given surfactant therapy on a regular basis due to the non-availability of effective low cost surfactant replacements. Hence, the need to develop a surfactant replacement which avoids the drawbacks of animal derived surfactant and the need for an effective synthetic surfactant replacement composition for ARDS has been felt for sometime.
OBJECT OF INVENTION
The object of this invention is to provide an exogenous protein free pulmonary surfactant composition, which is effective even in the presence of inhibitory
4

agents like plasma proteins and red cell membranes and which does not exhibit any harmful side effects and is also cost effective.
DESCRIPTION OF INVENTION and PREFRRED EMBODIMENTS
This invention relates to an exogenous protein free pulmonary surfactant composition for treating adult respiratory syndrome which comprises atleast one phospholipid surfactant and a natural oil selected from eucalyptus oil or its constituents.
This invention also includes a process for producing an exogenous protein free pulmonary surfactant composition for treating adult respiratory syndrome which comprises the steps of preparing a solution of atleast one phospholipid surfactant in an organic solvent, drying the said solution under vacuum, hydrating the residue thus obtained with aqueous isotonic electrolyte solutions and adding natural oils selected from eucalyptus oil or its constituents thereto.
Phospholipid surfactant capable of being used in this composition is dipalmitoyl phosphatidyl choline (PC).
The pH of the solution may be adjusted to 7 to 7.4 and the preferred organic solvent is chloroform : methanol (2:1).
5

Preferred composition may have 0.1 to 1% w/v of eucalyptus oil or its constituents.
Phospholipid natural oil ratio may be 70:30 to 10:90 w/v lipids with final concentrations in the range of 0.1 to 1%. Eucalyptus oil used in the range of 5 to 6 microlites.
Though eucalyptus oil is known to reduce phlegm in traditional medicines, it has not been used as a surfactant. None of the protein free surfactants can prevent interactions of the surfactants with the hematological agents. The uniqueness of this composition is that it contains one or two phospholipid constituents that are present in natural living surfactants which in the presence of eucalyptus oil or its constituents, prevents the interaction between surfactant and the hematological agents and hence overcomes inhibition achieving a function which is as good as natural surfactants in the absence of inhibitors.
Efficacy of this composition has been tested and is found to be better than the hitherto known artificial surfactants. Surfactants compositions of this invention containing eucalyptus oil are as effective in the presence of hematological inhibitors as animal protein based surfactants are in their absence and are hence
6

of benefit. Further, they are without the harmful side effects of animal proteins. Cost factor is also an important aspect of this invention.
Parameters for a surfactant to be effective for ARDS are (1) preventing interaction between surfactant and hematological agents (2) achievement of low minimum surface tension in the presence of hematological agents (3) low compressibility in the presence of hematological agents (4) requirement of a minimal area change for achievement of low surface tension in the presence of hematological agents. The composition of the subject invention fulfills all these conditions.
The following examples are only illustrative and are not intended to limit the scope of this invention.
Example 1:
A solution of dipalmitoyl phosphatidyl choline in chloroform: methanol (2:1) in the concentration of 10 mg / ml was allowed to evaporate. To the residue 1 mL of normal saline was added and vigorously shaken. Eucalyptus oil 6 microl was then added to this solution. The solution was left undisturbed to allow hydration for a short period example 1 hr. The final phospholipids oil concentration in saline was 0.1%.
7

Example 2:
The procedure of example 1 was followed using 12 hour hydration instead of 1 hour. Evaporation is performed to get a uniform thin film of PLS which facilitates uniform distribution of PLS in normal saline. Solvent in which PLS is to be dissolved may be chloroform, a mixture of chloroform and methanol in the ratio of 2:1. Other organic solutions may also be used in combination with methanol and / or chloroform. The hydration period controls the particle sizes of the formulations. Short periods of hydration for example 1 hour results in nanoparticles of 500-800 nm size which can be used in an inhalable form whereas longer periods of hydration for example 12 hours results in particles of a few microns which can be used as injections into the airways (intratracheal instillation).
Any isotonic polyelectrolyte solution may be used for hydration. The polyelectrolytes should not contain calcium to prevent adverse interactions with hematological agents in ARDS.
This invention described hereinabove and claimed in the appended claims do not exclude obvious equivalents, modifications and alternations.
8

WE CLAIM :-
1. An exogenous protein-free pulmonary surfactant composition for treating Adult Respiratory Syndrome comprising atleast one phospholipid surfactants and a natural oil selected from eucalyptus oil and/or its constituents.
2. The composition as claimed in claim 1 wherein the said composition is in the form of nanoparticles.
3. The composition as claimed in claim 1 wherein normal saline is used as a medium.
4. The composition as claimed in claims 1 to 3 wherein said phospholipid surfactant is dipalmitoyl phospahtidyl choline (PC).
5. The composition as claimed in claims 1 to 4 which contains 0.1 to 1% w/v of eucalyptus oil.
6. The composition as claimed in claims 1 to 5 wherein surface tension of composition on film compression in the presence of hematological agent is 0-5 mN/m.
7. A process for producing an exogenous protein-free pulmonary surfactant composition for treating adult respiratory syndrome comprising the steps of preparing a solution of atleast one phospholipid surfactants in an organic solvent, drying the same under vacuum, hydrating the residue obtained thereby with aqueous isotonic electrolyte solution and adding a natural oil selected from eucalyptus oil or its constituents thereto
9

The process as claimed in claim 7 wherein prior to hydration, the pH of
the solution is adjusted to 7.2 to 7.4
The process as claimed in claims 7-8 wherein said composition has a
phospholipid oil ratio of 70:30 to 10:90 w/v.
An exogenous protein free pulmonary surfactant composition for treating
adult respiratory syndrome substantially as herein described.
A process for preparing exogenous protein-free surfactant composition
for treating adult respiratory syndrome substantially as herein described
Dated this 30th day of September 2005

10

ABSTRACT
This invention relates to an exogenous protein-free surfactant composition for use in treating adult respiratory distress syndrome (ARDS) a process for preparing the surfactant composition. The composition contains alteast one phospholipid surfactant and a natural oil selected from eucalyptus oil and its derivatives. This composition prevents the interaction between surfactants and hematological agents and will thus be effective in ARDS. It is also cost effective and does not have the side -effects normally seen when protein containing surfactants are used.

Documents:

1223-MUM-2005-ABSTRACT(15-1-2010).pdf

1223-mum-2005-abstract(granted)-(25-6-2010).pdf

1223-mum-2005-abstract.doc

1223-mum-2005-abstract.pdf

1223-MUM-2005-CANCELLED PAGES(15-1-2010).pdf

1223-MUM-2005-CANCELLED PAGES(23-2-2010).pdf

1223-MUM-2005-CLAIMS(AMENDED)-(15-1-2010).pdf

1223-MUM-2005-CLAIMS(AMENDED)-(23-2-2010).pdf

1223-mum-2005-claims(granted)-(25-6-2010).pdf

1223-mum-2005-claims.doc

1223-mum-2005-claims.pdf

1223-mum-2005-correspondence(27-2-2008).pdf

1223-mum-2005-correspondence(ipo)-(16-2-2009).pdf

1223-mum-2005-correspondence(ipo)-(7-7-2010).pdf

1223-mum-2005-correspondence-received.pdf

1223-mum-2005-description (complete).pdf

1223-mum-2005-description(granted)-(25-6-2010).pdf

1223-MUM-2005-DRAWING(15-1-2010).pdf

1223-MUM-2005-DRAWING(23-2-2010).pdf

1223-mum-2005-drawing(granted)-(25-6-2010).pdf

1223-mum-2005-form 13(15-1-2010).pdf

1223-mum-2005-form 18(27-2-2008).pdf

1223-mum-2005-form 2(granted)-(25-6-2010).pdf

1223-mum-2005-form 2(title page)-(granted)-(25-6-2010).pdf

1223-MUM-2005-FORM 8(15-1-2010).pdf

1223-mum-2005-form-1.pdf

1223-mum-2005-form-2.doc

1223-mum-2005-form-2.pdf

1223-mum-2005-form-26.pdf

1223-mum-2005-form-3.pdf

1223-mum-2005-form-5.pdf

1223-mum-2005-marked copy(15-1-2010).pdf

1223-MUM-2005-REPLY TO EXAMINATION REPORT(15-1-2010).pdf

1223-MUM-2005-REPLY TO EXAMINATION REPORT(23-2-2010).pdf

1223-MUM-2005-SPECIFICATION(AMENDED)-(15-1-2010).pdf

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Patent Number

241272

Indian Patent Application Number

1223/MUM/2005

PG Journal Number

27/2010

Publication Date

02-Jul-2010

Grant Date

25-Jun-2010

Date of Filing

30-Sep-2005

Name of Patentee

INDIAN INSTITUTE OF TECHNOLOGY

Applicant Address

Powai, Mumbai 400 076,

Inventors:

#	Inventor's Name	Inventor's Address
1	RINTI BANERJEE	Powai, Mumbai 400 076,
2	RACHANA	Powai, Mumbai 400 076
3	JAYESH BELLARE	Powai, Mumbai 400 076
4	RAM PUNIYANI	Powai, Mumbai 400 076

PCT International Classification Number

C07H21/00

PCT International Application Number

N/A

PCT International Filing date

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1			NA