Drug – bio-affecting and body treating compositions – Preparations characterized by special physical form – Liposomes
Reexamination Certificate
1999-02-18
2003-11-11
Kishore, Gollamudi S. (Department: 1615)
Drug, bio-affecting and body treating compositions
Preparations characterized by special physical form
Liposomes
C514S021800
Reexamination Certificate
active
06645522
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a liposome based formulation of erythropoietin. In particular, the invention relates to a liposome based parenteral dosage form of erythropoietin prepared by means of an ethanol injection technique which exhibits superior stability.
BACKGROUND OF THE INVENTION
Erythropoietin (EPO) is a glycoprotein which serves as the principal factor involved in the regulation of red blood cell synthesis. Erythropoietin is produced in the kidney and acts by stimulating precursor cells in the bone marrow causing them to divide and differentiate into mature red blood cells. The recombinantly produced 165 amino acid glycoprotein has been available for some time as an effective therapeutic agent in the treatment of various forms of anemia, including anemias associated with chronic renal failure, zidovidine treated HIV infected patients, and cancer patients on chemotherapy. The glycoprotein is administered parenterally, either as an intravenous (IV) or subcutaneous (SC) injection.
Presently, the parenteral formulations in use are conventional sterile buffered aqueous solutions for IV or SC injection which contain human serum albumin (HSA) as a carrier. Such formulations are marketed in the United States under the trade names EPOGEN® and PROCRIT®. These products contain erythropoietin in 1 ml single dose, preservative-free or 2 ml multidose preserved vials..
While these formulations have been proven to be highly successful, certain disadvantages are associated with the use of human serum albumin as carrier. As HSA is obtained from natural sources it can be a potential danger as a carrier for infectious disease agents such as HIV or hepatitis and careful screening of the material must be conducted. Further, the availability of appropriate quality of HSA can often be a problem. Hence, there is a need for an injectable formulation of erythropoietin which eliminates the use of HSA as a carrier.
Accordingly, attempts have been made to provide an improved formulation of erythropoietin which eliminates the use of HSA as a carier. At the same time the formulation should be stable and provide an extended shelf life. Further, the formulation must avoid problems associated with the active ingredient adhering to the surface of the vial in which it is contained.
Liposomes are small vesicles comprising amphipathic lipids arranged in spherical bilayers. Liposomes may contain many concentric lipid bilayers separated by aqueous channels (multilamellar vesicles or MLVs), or alternatively, they may contain a single membrane bilayer (unilamellar vesicles), which may be small unilamellar vesicles (SUVs) or large unilamellar vesicles (LUVs). The lipid bilayer is composed of two lipid monolayers having a hydrophobic “tail” region and a hydrophilic “head” region. In the membrane bilayer, the hydrophobic “tails” of the lipid monolayers orient towards the center of the bilayer, whereas the hydrophilic “heads” orient toward the aqueous phase.
Liposomes may be used to encapsulate a variety of materials by trapping hydrophilic compounds in the aqueous interior or between bilayers, or by trapping hydrophobic compounds within the bilayer. As such, they are particularly useful to deliver biologically active materials by encapsulating compounds which exhibit poor aqueous solubility or which exhibit unacceptable toxicity at therapeutic dosages.
A specific method for the production of liposomes with only one double layer is disclosed in EP 253 619. Liposome formulations of various active agents have been known for years and liposomal preparations of erythropoietin have been proposed. For example, Maitani et al, J. Pharm. Sci., 85:440-445 (1996) discloses liposomal erythropoietin formulations intended for oral administration in which the liposomes are prepared by the reverse phase evaporation vehicle method. Since the formulation therein is intended for oral administration, a high percentage of incorporation of EPO into the liposomes is preferred. However, formulations such as this demonstrating a high rate of encapsulation in small vesicles may exhibit concentration in the liver, leading to toxicities. Moreover, the manufacturing procedures used therein require special raw materials (e.g. polyglycerine phospholipid) and the use of organic solvents. Further, the reverse phase process used therein suffers a high loss of unencapsulated EPO, which is undesirable and expensive.
The goal of the present invention therefore was to provide a parenteral formulation suitable for EPO, which avoids the use of HSA as carrier, provides acceptable long term stability for an extended shelf life, and which can be manufactured by means of a process which is amenable to large scale manufacture.
SUMMARY OF THE INVENTION
A liposome-based parenteral composition comprising:
(a) an effective amount of an active ingredient comprising erythropoietin or its pharmaceutically acceptable derivatives having the biological properties of causing bone marrow cells to increase production of reticulocytes and red blood cells;
(b) a lipidic phase comprising:
(i) lecithin or hydrogenated lecithin;
(ii) optionally, a charged electropositive or electronegative lipid compound; and
(iii) cholesterol or a derivative thereof selected from cholesterol esters, polyethylene glycol derivatives of cholesterol (PEG-cholesterols), and organic acid derivatives of cholesterols; and
(c) an aqueous buffer solution.
In accordance with the invention, the composition comprises single bilayered liposomes made by preparing an alcoholic solution of the lipidic phase and injecting the solution under pressure into an aqueous buffer solution contained in a high speed homogenizer. The liposomes thus prepared are incubated with the erythropoietin active ingredient to form the liposomal dispersion of the invention.
Preferably, the active ingredient is erythropoietin and its derivatives having the biological properties of causing bone marrow cells to increase production of reticulocytes and red blood cells. The EPO glycoprotein may be obtained from natural sources or produced recombinantly using known procedures as disclosed in U.S. Pat. Nos. 4,703,008, 5,441,868, 5,547,933, 5,618,698 and 5,621,080, hereby incorporated by reference.
In accordance with the present invention, it has been discovered that, quite unexpectedly, the liposomal EPO compositions prepared under the mild conditions described herein exhibit improved stability, i.e. the liposomes themselves are stable and at the same time the chemical degradation and aggregation of the biologically effective substance is minimized. As a further unexpected advantage, the EPO active ingredient does not adhere to the surface of the vial container or IV tubing even though the EPO is not substantially incorporated within the liposomes, but is instead essentially contained in the interstitial fluid as a liposomal dispersion.
DETAILED DESCRIPTION
The active ingredient used in the present invention is erythropoietin and its derivatives having the biological properties of causing bone marrow cells to increase production of reticulocytes and red blood cells. The liposomal dispersion of the present invention is useful as a parenteral formulation in treating blood disorders characterized by low or defective red blood cell production such as various forms of anemia, including anemias associated with chronic renal failure, zidovidine treated HIV infected patients, and cancer patients on chemotherapy. It may also have application in the treatment of a variety of disease states, disorders and states of hematologic irregularity such as sickle cell disease, beta-thalassemia, cystic fibrosis, pregnancy and menstrual disorders, early anemia of prematurity, spinal cord injury, space flight, acute blood loss, aging and the like. Preferably, the EPO composition of the present invention is administered parenterally (e.g. IV, IM, SC or IP). Effective dosages are expected to vary considerably depending on the condition being treated and the route of administration but are expected to be in the range of 0.1 (~7U) to 1
Delmenico Sandro
Floether Frank-Ulrich
Naeff Rainer
Wetter André
Cilag AG
Kentoffio Joseph S.
Kishore Gollamudi S.
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