Title of Invention

A NANOPARTICULATE EXOGENOUS PROTEIN-FREE MICRONUTRIENT ADDED PULMONARY SURFACTANT COMPOSITION FOR NEONATAL RESPIRATORY DISTRESS SYNDROME AND A PROCESS FOR PREPARING THE SAME.

Abstract This invention relates to a nanoparticulate exogenous micronutrient added protein-free surface composition for use in treating Neonatal Respiratory Distress Syndrome (NRDS) and a process for preparing the same. The composition contains atleast one phospholipid surfactant and a natural oil selected from clove or eucalyptus oil and their derivatives, micronutrients selected from ascorbic acid and its derivatives and soluble calcium salts. This composition is cost effective, does not have the side -effects normally seen when animal derived protein containing surfactants are used and acts also as a delivery agent for essential micronutrients for the premature babies lungs.
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
A nanoparticulate exogenous protein-free micronutrient
added pulmonary surfactant compostion for
neonatal 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 micronutrient added pulmonary surfactant composition useful in treating respiratory disorders like Neonatal Respiratory Distress Syndrome (NRDS) which are due to a deficiency of pulmonary surfactant and a process for preparing such surfactant composition.
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, phosphatidyl glycerol, unsaturated lecithins, cholesterol, surfactant specific proteins, phosphatidyl ethanolamine, glycerides, phosphatidyl serine, lysolecithin and fatty acids. Due to deficiency 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 in order to re-expand the alveoli with each intake of breath. This condition is recognized as neonatal respiratory distress syndrome (NRDS) and occurs in premature infants, born before their lungs can synthesize surfactant.
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Presently infants having Neonatal Respiratory Distress Syndrome are provided with intensive care in incubators with positive pressure mechanical ventilation with oxygen supplementation. However, this treatment only has temporary effect and does not correct the underlying pathogenic cause.
Surfactant replacement therapy has been found beneficial in treating NRDS, asthma and other chronic bronchial problems. Exogenous surfactants as replacement or supplement have been developed and are in use for over three decades in developed countries for NRDS. Many of these formulations contain animal derived proteins and are obtained mainly from cows, pigs and goats. Animal derived synthetic surfactant compositions are highly efficient. 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 for developing countries like India.
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 polyelectrolyte buffers. Compositions having PC in combination with co-
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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, higher values of surface tension and compressibility, as well as a poor respreading ratio. These properties limit their effectiveness as a therapy in NRDS.
NRDS patients in India are not given surfactants therapy on a regular basis due to the non-availability of effective low cost surfactant replacements. Imported surfactants are not affordable to the majority of Indian population. Hence, the need to develop a surfactant replacement which avoids the drawbacks of animal derived surfactants and the hitherto need for a synthetic surfactant replacement composition has been felt for sometime.
Further, premature babies lack several nutrients in adequate quantities requiring their supplementation. One such constituent is vitamin C or ascorbic acid, which is an essential micronutrient for lung development and has anti-oxidant properties. Also, there is a need to deliver the surfactants into the deep portions of the lungs (alveoli) in a non-invasive way. One way to achieve this is by the use of nanoparticles which can reach the alveoli when used in an inhalable form. The present surfactants have larger several micron sized particles and are delivered by injections into the airways (intratracheal bolus injections) which are very invasive.
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OBJECT OF INVENTION
The object of this invention is to provide an exogenous protein-free pulmonary surfactant composition which is affordable to the common man, which is as effective as the animal derived surfactants, which contains micronutrients (thus acting as a delivery agent for micronutrients in the lungs), which can be given in a non-invasive inhalable form and which does not exhibit any harmful side effects as it lacks animal proteins.
DESCRIPTION OF INVENTION and PREFERRED EMBODIMENTS
This invention relates to a nanoparticulate exogenous protein-free pulmonary surfactant composition for treating neonatal respiratory distress syndrome which comprises atleast one phospholipid surfactant and a natural oil selected from clove oil, or its constituents and eucalyptus oil and its derivatives or constituents and micronutrients selected from ascorbic acid and a soluble calcium salt either alone or in combination.
This invention also includes a process for producing a nano-particulate exogenous protein free pulmonary surfactant composition for treating neonatal respiratory distress syndrome which comprises the steps of preparing a solution of atleast one phospholipid surfactant in an organic solvent, drying the said solution under vacuum, adding an aqueous isotonic electrolyte solution
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containing micronutrients selected from soluble calcium salt, adding ascorbic acid and natural oils selected from clove oil or eucalyptus oil thereto and allowing the residue to hydrate.
Phospholipid surfactants capable of being used in this composition are dipalmetoyl phosphatidyl choline (PC), Phosphatidyl ethanolamine (PE) and /or phosphatidyl glycerol (PG). In a preferred embodiment where more than one phospholipid surfactant are used their ratio may be PC to PE or PC to PG = 2:3
The pH of the solution may be adjusted to 7 to 7.4 and the preferred organic solvent is chloroform.
Preferred composition may have 0.1 to 1% w/v of eucalyptus oil or about 1% by clove oil.
Preferred composition has particle size 150 to 450 nm in diameters.
Phospholipid natural oil ratio may be 70:30 to 10:90 w/v with final concentrations in the range of 0.1 to 1% and clove oil and eucalyptus oil are used in the range of 5 to 6 microlites.
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Clove oil improves respiratory dynamics and is an emulsifying agent. Though eucalyptus oil is known to reduce phlegm in traditional medicines, it has not been used as a surfactant. 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 either clove oil or eucalyptus oil achieves a function which is as good as natural surfactants. The short hydration period, for example 1 hour hydration, ensures the formation of nanoparticles that can be inhaled into the lungs easily. The PLS and clove oil or eucalyptus oil combination will help in the uniform dispersion of ascorbic acid into the lungs and act as a delivery agent for this micronutrient cum antioxidant.
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 clove or eucalyptus oil and ascorbic acid in the presence of calcium salts are as effective as animal protein based surfactants but are without their harmful side effects. Cost factor is also an important aspect of this invention.
Main parameters for a surfactant to be effective in NRDS are (1) minimum surface tension on film compression 7

inhalation (as a nebulization) and (5) uniform delivery of micronutrients essential for the premature lungs.
The following examples are only illustrative and are not intended to limit the scope of this invention.
Example 1:
A 1 ml 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 Ringer's lactate containing 2 mM calcium chloride was added and vigorously shaken. Ascorbic acid was added in the concentration of 0.1 mg/ml to this solution. Clove oil 1% w/v was then added to this solution was left undisturbed and allowed to hydrate for short periods for example 1 hr.
Example 2 :
The procedure of example 1 was followed using a mixture of dipalmitoyl phosphatidyl choline and phosphatidyl ethanolamine
Example 3:
The procedure of examples 1 and 2 is repeated with eucalyptus oil instead of clove oil.
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Example 4:
The procedure of examples 1 and 2 is repeated with 12 hours hydration instead of 1 hour and extrusion of the surfactant post-hydration through lOOnm pore sized polycarbonate filters.
Evaporation is performed to get a uniform thin film of PLS which facilitates its uniform distribution in Ringer's solution. 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.
Any isotonic polyelectrolyte solution may be used for hydration. Polyelectrolyte with calcium is preferred for use in NRDS.
This invention described hereinabove and claimed in the appended claims do not exclude obvious equivalents, modifications and alternations.


WE CLAIM :
1. A nanoparticulate exogenous protein-free pulmonary surfactant composition for treating Neonatal Respiratory Distress Syndrome comprising atleast one phospholipid surfactants and a natural oil selected form clove oil and its constituents and/or eucalyptus oil and or its constituents with micronutrients selected from ascorbic acid and soluble calcium salts either alone or in combination.
2. The composition as claimed in claim 1 wherein ascorbic acid is preferably 0.1 mg/ml.
3. The composition as claimed in claims 1 - 2 wherein particle size is preferably in 150 - 450 nm in diameter.
4. The composition as claimed in claims 1 - 3 wherein surface tension of composition on film compression is 0 - 5mN/m.
5. The composition as claimed in claims 1 - 4 wherein said phospholipid surfactants are dipalmitoyl phospahtidyl choline (PC), phosphatidyl elhanolamine (PE) and/or phosphatidyl glycerol (PG) or mixtures thereof.
6. The composition as claimed in claim 3 wherein the ratio of PC to PE or PE to PG is 2:3.
7. The composition as claimed in claim 1 which contains either ascorbic acid or soluble calcium salt or both.
8. The composition as claimed in claims 1-6 which contains 0.1 to 1% w/v of eucalyptus oil or about 1% of clove oil.
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9. A process for producing a nanoparticulate exogenous protein-free pulmonary surfactant composition for treating Neonatal Respiratory Distress Syndrome comprising the steps of preparing a solution of atleast one phospholipid surfactants in an organic solvent, drying the same under vacuum, adding aqueous isotonic electrolyte solution, adding a natural oil selected from clove oil or eucalyptus oil thereto, micronutrients selected from ascorbic acid and its derivatives and soluble calcium salts, either alone or in combination and allowing the mixture to hydrate.
10^- The process as claimed in claim 9 wherein the pH of the solution is adjusted to 7.2 to 7.4 prior to hydration.
11. The process as claimed in claim 9 wherein said composition has a phospholipid oil ratio of 70:30 to 10:90 w/v
12. The process as claimed in claim 9 wherein the hydration period is about 1 hour and/or 12 hours thereafter surfactant is extruded through submicron filters.
13. A nanoparticulate exogenous protein-free micronutrient surfactant composition for treating Neonatal Respiratory Distress Syndrome substantially as herein described.
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14. A process for preparing nanoparticulate exogenous protein-free micronutrient composition for treating NRDS substantially as herein described.
Dated this 30th day of September 2005

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ABSTRACT
This invention relates to a nanoparticulate exogenous micronutrient added protein-free surfactant composition for use in treating Neonatal Respiratory Distress Syndrome (NRDS) and a process for preparing the same. The composition contains alteast one phospholipid surfactant and a natural oil selected from clove or eucalyptus oil and their derivatives, micronutrients selected from ascorbic acid and its derivatives and soluble calcium salts. This composition is cost effective, does not have the side - effects normally seen when animal derived protein containing surfactants are used and acts also as a delivery agent for essential micronutrients for the premature babies' lungs.

Documents:

1222-MUM-2005-ABSTRACT(15-1-2010).pdf

1222-mum-2005-abstract(granted)-(25-6-2010).pdf

1222-mum-2005-abstract.doc

1222-mum-2005-abstract.pdf

1222-MUM-2005-CANCELLED PAGES(15-1-2010).pdf

1222-mum-2005-cancelled pages(23-2-2010).pdf

1222-MUM-2005-CLAIMS(AMENDED)-(15-1-2010).pdf

1222-MUM-2005-CLAIMS(AMENDED)-(23-2-2010).pdf

1222-mum-2005-claims(granted)-(25-6-2010).pdf

1222-mum-2005-claims.doc

1222-mum-2005-claims.pdf

1222-mum-2005-correspondenc(27-2-2008).pdf

1222-mum-2005-correspondenc(ipo)-(12-2-2009).pdf

1222-MUM-2005-CORRESPONDENCE(23-2-2010).pdf

1222-mum-2005-correspondence(ipo)-(7-7-2010).pdf

1222-mum-2005-correspondence-received.pdf

1222-mum-2005-description (complete).pdf

1222-mum-2005-description(granted)-(25-6-2010).pdf

1222-MUM-2005-DRAWING(15-1-2010).pdf

1222-MUM-2005-DRAWING(23-2-2010).pdf

1222-mum-2005-drawing(granted)-(25-6-2010).pdf

1222-mum-2005-form 13(15-1-2010).pdf

1222-mum-2005-form 18(27-2-2008).pdf

1222-mum-2005-form 2(granted)-(25-6-2010).pdf

1222-mum-2005-form 2(title page)-(granted)-(25-6-2010).pdf

1222-MUM-2005-FORM 8(15-1-2010).pdf

1222-mum-2005-form-1.pdf

1222-mum-2005-form-2.doc

1222-mum-2005-form-2.pdf

1222-mum-2005-form-26.pdf

1222-mum-2005-form-3.pdf

1222-mum-2005-form-5.pdf

1222-mum-2005-marked copy(23-2-2010).pdf

1222-MUM-2005-REPLY TO EXAMINATION REPORT(15-1-2010).pdf

1222-MUM-2005-SPECIFICATION(AMENDED)-(15-1-2010).pdf

1222-MUM-2005-SPECIFICATION(AMENDED)-(23-2-2010).pdf


Patent Number 241271
Indian Patent Application Number 1222/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, Bombay
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 A61K45/06
PCT International Application Number N/A
PCT International Filing date
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 NA