Thrombopoietin compositions

Drug – bio-affecting and body treating compositions – Lymphokine

Reexamination Certificate

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Reexamination Certificate

active

06790439

ABSTRACT:

BACKGROUND OF THE INVENTION
Hematopoiesis is the process by which blood cells develop and differentiate from pluripotent stem cells in the bone marrow. This process involves a complex interplay of polypeptide growth factors (cytokines) acting via membrane-bound receptors on the target cells. Cytokine action results in cellular proliferation and differentiation, with a response to a particular cytokine often being lineage-specific and/or stage-specific. Development of a single cell type, such as a platelet, from a stem cell may require the coordinated action of a plurality of cytokines acting in the proper sequence.
Various cytokines have been developed as therapeutic agents. For example, erythropoietin, which stimulates the development of erythrocytes, is used in the treatment of anemia arising from renal failure. Several of the colony stimulating factors have been used in conjunction with cancer chemotherapy to speed the recovery of patients' immune systems. Interleukin-2, &agr;-interferon and &ggr;-interferon are used in the treatment of certain cancers.
An activity that stimulates megakaryocytopoiesis and thrombocytopoiesis has been identified in body fluids of thrombocytopenic animals and is referred to in the literature as “thrombopoietin” (recently reviewed by McDonald, Exp.
Hematol
. 16:201-205, 1988 and McDonald,
Am. J. Ped. Hematol. Oncol
. 14:8-21, 1992). This protein has now been produced using genetically engineered cultured cells. See, de Sauvage et al.,
Nature
369:533-538, 1994; Lok et al.,
Nature
369:565-568, 1994; Kaushansky et al.,
Nature
369:568-571, 1994; and Bartley et al.,
Cell
77:1117-1124, 1994.
Human thrombopoietin (TPO) is a 70 kD glycoprotein of 332 amino acid residues. It contains ah amino-terminal growth factor domain of approximately 152 residues and a carboxyl-terminal domain rich in carbohydrate. Truncated forms of TPO comprising the amino-terminal domain exhibit biological activity in vitro and in vivo. Non-human TPOs have also been described in the scientific literature (e.g., Lok et al., ibid.; Bartley et al., ibid.; Shimada et al.,
Blood
84 (10 Suppl. 1):326a, 1994).
Thrombopoietin appears to be subject to proteolysis and was isolated in heterogeneous or degraded form (Bartley et al., ibid.; de Sauvage et al., ibid.). Preparations of thrbmbopoietin reported in the scientific literature are therefore not well characterized as to composition and the relative activities of the various molecular species, although at least some of the proteolytic products are biologically active. However, little work has been reported to date on the large-scale production of thrombopoietin, and there is a need in the art for compositions of TPO that are suitable for pharmaceutical use. Such formulations should be stable on storage and easy to use.
SUMMARY OF THE INVENTION
It is an object of the present invention to provide pharmaceutical compositions of TPO, including aqueous compositions, that are stable on storage.
It is a further object of the invention to provide methods of reducing adsorption of TPO to surfaces, including surfaces of storage vials and filters used in the preparation and packaging of pharmaceutical compositions of TPO.
It is an additional object of the invention to provide methods for stabilizing pharmaceutical compositions of TPO, including aqueous solutions and lyophilized powders.
In one aspect, the present invention provides a composition comprising TPO, a physiologically acceptable buffer, a surface adsorption inhibitor selected from the group consisting of non-ionic surfactants and polyols, and an isotonic amount of a physiologically acceptable salt. The TPO may be human TPO or non-human (e.g., rat, mouse, dog or non-human primate) TPO. Within certain embodiments of the invention, the composition is an aqueous solution having a pH of 5.0 to 7.0, preferably 5.5 to 6.5. Within an alternative embodiment, the composition is a lyophilized powder. Within a further embodiment, the composition comprises histidine in an amount sufficient to reduce aggregation of the thrombopoietin. Within another embodiment, the composition is an aqueous pharmaceutical composition comprising thrombopoietin, 10-100 mM phosphate buffer, 0.01-1.0% polysorbate 20 or polysorbate 80, and an isotonic amount of sodium chloride, the composition having a pH of approximately 6.0.
Within another aspect, the present invention provides a method for reducing adsorption of thrombopoietin to a surface comprising adding to an aqueous solution of thrombopoietin an effective amount of a surface adsorption inhibitor selected from the group consisting of non-ionic surfactants and polyols prior to contacting the solution with the surface. Within selected embodiments of the invention the surface adsorption inhibitor is a polyoxyethylene sorbitan fatty acid ester, such as polysorbate 20 or polysorbate 80.
Within a third aspect, the present invention provides a method of stabilizing a composition of thrombopoietin comprising adding to the composition an effective amount of histidine.
These and other aspects of the invention will become evident upon reference to the following detailed description and the attached drawings.


REFERENCES:
patent: 4637834 (1987-01-01), Thurow
patent: 5096885 (1992-03-01), Pearlman et al.
patent: 5344644 (1994-09-01), Igari et al.
patent: 5358708 (1994-10-01), Patel
patent: 5534269 (1996-07-01), Igari et al.
patent: 5580856 (1996-12-01), Prestrelski et al.
patent: 94/03198 (1994-02-01), None
Manning et al., Pharm. Res., vol. 6(11), pp. 903 to 918, 1989.*
McDonald et al.,Exp. Hematol. 17: 865-871, 1989.
Hill et al.,Exp. Hematol. 17: 903-907, 1989.
McDonald,Exp. Hematol. 16: 201-205, 1988.
Foster et al.,Proc. Natl. Acad. Sci. U.S.A. 91: 13023-13027, 1994.
Wang et al.,Journal of the Parenteral Drug Association 34: 452-462, 1980.

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