Patentaml
`European
`Patent Office
`
`
`
`(m
`
`EP1 528 933 B1
`
`(12)
`
`EUROPEAN PATENT SPECIFICATION
`
`(45) Date of publication and mention
`of the grant of the patent:
`02.05.2012 Bulletin 2012/18
`
`(51)
`
`Int CI.:
`A61K 39/395 (Em-01)
`461K 47/26“"”""”
`
`A61K 9/19(2005.a1)
`
`(21) Application number: 03748439.1
`
`(85)
`
`International application number:
`PCT/lB2003/004502
`
`(22) Date of filing: 15.08.2003
`
`(87)
`
`International publication number:
`WO 2004/016286 (26.02.2004 Gazette 2004/09)
`
`(54) PHARMACEUTICAL ANTI-TNF-ALPHA ANTIBODY FORMULATION
`
`PHARMEZEUTISCHE ANTI-TNF-ALPHA ANTIKORPER FORMULIERUNG
`
`FORMULATION PHARMACEUTIQUE D’ANTICORPS ANTI-TNF-ALPHA
`
`(84) Designated Contracting States:
`AT BE BG CH CY CZ DE DK EE ES FI FR GB GR
`
`US-A- 6 090 382
`
`US-B1- 6 171 586
`
`HU IE IT LI LU MC NL PT RO SE SI SK TR
`
`(30) Priority: 16.08.2002 US 222140
`
`(43) Date of publication of application:
`11.05.2005 Bulletin 2005/19
`
`(60) Divisional application:
`101825982 /2 363 144
`101826105 /2 359 856
`
`101826196 / 2 361 637
`101826253 / 2 363 145
`
`(73) Proprietor: Abbott Biotechnology Ltd.
`HM 11 Hamilton (BM)
`
`Inventors:
`(72)
`0 KRAUSE, Hans-Juergen
`68647 Biblis (DE)
`o BAUST, Lisa
`67063 Ludwigshafen (DE)
`0 DICKES, Michael
`67127 Rodersheim-Gronau (DE)
`
`(74) Representative: Reitst6tter - Kinzebach
`Patentanwalte
`
`Sternwartstrasse 4
`81679 Miinchen (DE)
`
`(56) References cited:
`EP-A- 1 174 148 WO-A-01/47554
`WO-A—02/12502 WO-A-97/04801
`WO-A—98/5641 8 WO-A1-97I29131
`US-A- 5 945 098
`US-A— 6 024 938
`
`0 CLELAND JEFFREY L ET AL: "A specific molar
`ratio of stabilizerto protein is required for storage
`stability of a lyophilized monoclonal antibody"
`JOURNAL OF PHARMACEUTICAL SCIENCES,
`vol. 90, no. 3, March 2001 (2001-03), pages
`310-321, XP002278944 ISSN: 0022-3549
`0 HlLLGREN ANNA ET AL: "Protection mechanism
`
`of Tween 80 during freeze-thawing of a model
`protein, LDH." INTERNATIONAL JOURNAL OF
`PHARMACEUTICS. NETHERLANDS 26 APR
`
`2002, vol. 237, no. 1-2, 26 April 2002 (2002-04-26),
`pages 57-69, XP002278945 ISSN: 0378-5173
`0 CARPENTER-JF ET AL.: "Rational design of
`stable lyophilized protein formulations: some
`practical advice", PHARM RES, vol. 14, no. 8,
`August 1997 (1997-08), page 969-75,
`- WANG-W: "Instability, stabilization, and
`formulation of liquid protein pharmaceuticals",
`INT J PHARM, vol. 185, no. 2, 20 August 1999
`(1999-08-20), page 129-88, XP002616539,
`ISBN-10: 0306467453, 30 June 2002 (2002-06-30),
`in: Development and Manufacture of Protein
`Pharmaceuticals (Pharmaceutical
`Biotechnology)
`0 AKERS-MJ ET AL.: "Development and
`Manufacture of Protein Pharmaceuticals
`
`0
`
`(Pharmaceutical Biotechnology): chapter 2
`Formulation Development of Protein Dosage
`Forms", 30 June 2002 (2002-06-30), KLUWER
`ACADEMIC/PLENUM PUBLISHER, NEW YORK,
`XP001537612, * the whole document *
`
`
`
`Note: Within nine months of the publication of the mention of the grant of the European patent in the European Patent
`Bulletin, any person may give notice to the European Patent Office of opposition to that patent, in accordance with the
`Implementing Regulations. Notice of opposition shall not be deemed to have been filed until the opposition fee has been
`paid. (Art. 99(1) European Patent Convention).
`
`Printed by Jouve, 75001 PARIS (FR)
`
`EP1528933B1
`
`AMGEN INC.
`
`Exhibit 1052
`
`Ex. 1052 - Page 1 of 29
`
`Ex. 1052 - Page 1 of 29
`
`AMGEN INC.
`Exhibit 1052
`
`

`

`Europilsches
`Patentaml
`European
`Patent Office
`Office européen
`des brevets
`
`
`
`EUROPEAN PATENT SPECIFICATION
`
`(11)
`
`EP1 528 933 BI
`
`Date of publication and mention
`of the grant of the patent:
`02.05.2012 Bulletin 2012/18
`
`(51)
`
`Int CI.:
`A61K 39/395 (am-"1)
`A61K 47/26 (2mm)
`
`A61K 9/19 (2006-01)
`
`(12)
`
`(45)
`
`(21)
`
`Application number: 03748439.1
`
`(22)
`
`Date of filing: 15.08.2003
`
`(86)
`
`International application number:
`PCT/lB2003/004502
`
`(87)
`
`International publication number:
`WO 2004/016286 (26.02.2004 Gazette 2004/09)
`
`(54)
`
`PHARMACEUTICAL ANTI-TNF-ALPHA ANTIBODY FORMULATION
`
`PHARMEZEUTISCHE ANTI-TNF-ALPHA ANTIKORPER FORMULIERUNG
`
`FORMULATION PHARMACEUTIQUE D’ANTICORPS ANTI-TNF-ALPHA
`
`(84)
`
`Designated Contracting States:
`AT BE BG CH CY CZ DE DK EE ES FI FR GB GR
`HU IE IT LI LU MC NL PT RO SE SI SK TR
`
`(30)
`
`Priority: 16.08.2002 US 222140
`
`(43)
`
`Date of publication of application:
`11.05.2005 Bulletin 2005/19
`
`(50)
`
`Divisional application:
`101825982 / 2 363 144
`10182610.5 / 2 359 856
`10182619.6 / 2 361 637
`10182625.3 / 2 363 145
`
`(73)
`
`Proprietor: Abbott Biotechnology Ltd.
`HM 11 Hamilton (BM)
`
`Inventors:
`
`(72)
`
`KRAUSE, Hans-Juergen
`68647 Biblis (DE)
`BAUST, Lisa
`67063 Ludwigshafen (DE)
`DICKES, Michael
`67127 Rodersheim-Gronau (DE)
`
`(74)
`
`Representative: Reitst6tter - Kinzebach
`Patentanwalte
`Sternwartstrasse 4
`
`81679 Miinchen (DE)
`
`(56)
`
`References cited:
`EP-A- 1 174148
`WO-A-02/12502
`WO-A—98/5641 8
`US-A- 5 945 098
`
`WO-A-01/47554
`WO-A-97/04801
`WO-A1-97I29131
`US-A- 6 024 938
`
`US-A- 6 090 382
`
`US-B1- 6 171 586
`
`CLELAND JEFFREY L ET AL: "A specific molar
`ratio of stabilizerto protein is required for storage
`stability of a lyophilized monoclonal antibody"
`JOURNAL OF PHARMACEUTICAL SCIENCES,
`vol. 90, no. 3, March 2001 (2001-03), pages
`310-321, XP002278944 ISSN: 0022-3549
`HlLLGREN ANNA ET AL: "Protection mechanism
`
`of Tween 80 during freeze-thawing of a model
`protein, LDH." INTERNATIONAL JOURNAL OF
`PHARMACEUTICS. NETHERLANDS 26 APR
`
`2002, vol. 237, no. 1-2, 26 April 2002 (2002-04-26),
`pages 57-69, XP002278945 ISSN: 0378-5173
`CARPENTER-JF ET AL.: "Rational design of
`stable lyophilized protein formulations: some
`practical advice", PHARM RES, vol. 14, no. 8,
`August 1997 (1997-08), page 969-75,
`WANG-W: "Instability, stabilization, and
`formulation of liquid protein pharmaceuticals",
`INT J PHARM, vol. 185, no. 2, 20 August 1999
`(1999-08-20), page 129-88, XP002616539,
`ISBN-10: 0306467453, 30 June 2002 (2002-06-30),
`in: Development and Manufacture of Protein
`Pharmaceuticals (Pharmaceutical
`Biotechnology)
`AKERS-MJ ET AL.: " Development and
`Manufacture of Protein Pharmaceuticals
`
`(Pharmaceutical Biotechnology): chapter 2
`Formulation Development of Protein Dosage
`Forms", 30 June 2002 (2002-06-30), KLUWER
`ACADEMIC/PLENUM PUBLISHER, NEW YORK,
`XP001537612, * the whole document *
`
`
`
`Note: Within nine months of the publication of the mention of the grant of the European patent In the European Patent
`Bulletin, any person may give notice to the European Patent Office of opposition to that patent, in accordance with the
`Implementing Regulations. Notice of opposition shall not be deemed to have been filed until the opposition fee has been
`paid. (Art. 99(1) European Patent Convention).
`
`Printed by Jouve, 75001 PARIS (FR)
`
`(Cont. next page)
`
`EP1528933B1
`
`Ex. 1052 - Page 2 of 29
`
`Ex. 1052 - Page 2 of 29
`
`

`

`EP1 528 933 B1
`
`0 BARRERA P ET AL: "Effects of treatment with a
`fully human anti-tumour necrosis factor alpha
`monoclonal antibody on the local and systemic
`homeostasis of interleukin 1 and TNFalpha in
`patients with rheumatoid arthritis.", ANNALS OF
`THE RHEUMATIC DISEASES JUL 2001 LNKD-
`
`PUBMED:11406520, vol. 60, no. 7, July 2001
`(2001-07), pages 660-669, ISSN: 0003-4967
`0 HARRIS REED J ET AL: "Commercial
`manufacturing scale formulation and analytical
`characterization of therapeutic recombinant
`antibodies", DRUG DEVELOPMENT RESEARCH,
`vol. 61, no. 3, March 2004 (2004-03), pages
`137-154, ISSN: 0272-4391
`
`0 WANG WEI ET AL: "Antibody structure,
`instability, and formulation.", JOURNAL OF
`PHARMACEUTICAL SCIENCES JAN 2007 LN KD-
`PUBMED:16998873, vol. 96, no. 1, January 2007
`(2007-01), pages 1-26, ISSN: 0022-3549
`
`Remarks:
`Thefile containstechnioal information submitted after
`the application was filed and not included in this
`specification
`
`
`
`Ex. 1052 - Page 3 of 29
`
`Ex. 1052 - Page 3 of 29
`
`

`

`EP 1 528 933 B1
`
`Description
`
`Background of the Invention
`
`[0001] Tumor necrosis factor a (TNFa) is a cytokine produced by numerous cell types, including monocytes and
`macrophages, that was originally identified based on its capacity to induce the necrosis of certain mouse tumors (see
`e.g., Old, L. (1985) Science 230:630-632). Subsequently, a factor termed cachectin, associated with cachexia, was
`shown to be the same molecule as TNFa. TNFa has been implicated in mediating shock (see e.g., Beutler, B. and
`Cerami, A. (1988)Annu. Rev. Biochem. 57:505-518; Beutler, B. and Cerami, A. (1989)Annu. Rev. Immunol. 7:625-655).
`Furthermore, TNFoc has been implicated in the pathophysiology of a variety of other human diseases and disorders,
`including sepsis, infections, autoimmune diseases, transplant rejection and graft—versus—host disease (see e.g., Moeller,
`A., et al. (1990) Cytokine 2:162-169; U.S. Patent No. 5,231,024 to Moeller et al.; European Patent Publication No. 260
`610 B1 by Moeller, A., et al. Vasilli, P. (1992) Annu. Rev. Immunol. 10:411-452; Tracey, K.J. and Cerami, A. (1994)
`Annu. Rev. Med. 45:491—503).
`[0002] Because of the harmful role of human TNFoc (hTNFoz) in a variety of human disorders, therapeutic strategies
`have been designed to inhibit or counteract hTNFoc activity. In particular, antibodies that bind to, and neutralize, hTNFoc
`have been sought as a means to inhibit hTN Foe activity. Some of the earliest of such antibodies were mouse monoclonal
`antibodies (mAbs), secreted by hybridomas prepared from lymphocytes of mice immunized with hTN For (see e.g., Hahn
`T; et al., (1985) Proc Natl Acad Sci USA 82: 3814-3818; Liang, C-M., et al. (1986) Biochem. Biophys. Res. Commun.
`137:847-854; Hirai, M., et al. (1987) J. Immunol. Methods 96:57-62; Fendly, B.M., et al. (1987) Hybridoma 6:359—370;
`Moeller, A., et al. (1990) Cytokine 2:162-169; U.S. Patent No. 5,231,024 to Moeller et al.; European Patent Publication
`No. 186 833 B1 by Wallach, D.; European Patent Application Publication No. 218 868 A1 by Old et al.; European Patent
`Publication No. 260 610 B1 by Moeller, A., et al.). While these mouse anti-hTNFa antibodies often displayed high affinity
`for hTNFoc (e.g., Kd s 10'9M) and were able to neutralize hTNFa activity, their use in vivo may be limited by problems
`associated with administration of mouse antibodies to humans, such as short serum half life, an inabilityto trigger certain
`human effector functions and elicitation of an unwanted immune response against the mouse antibody in a human (the
`"human antimouse antibody" (HAMA) reaction).
`[0003]
`In an attempt to overcome the problems associated with use of fully-murine antibodies in humans, murine anti-
`hTNFoc antibodies have been genetically engineered to be more "human-like." For example, chimeric antibodies, in
`which the variable regions of the antibody chains are murine-derived and the constant regions of the antibody chains
`are human-derived, have been prepared (Knight, D.M, et al. (1993) Mol. Immunol. 30:1443-1453; PCT Publication No.
`WO 92/16553 by Daddona, P.E., et al.). Additionally, humanized antibodies, in which the hypen/ariable domains of the
`antibody variable regions are murine-derived butthe remainder ofthe variable regions and the antibody constant regions
`are human-derived, have also been prepared (PCT Publication No. WO 92/11383 by Adair, JR, et al.). However,
`because these chimeric and humanized antibodies still retain some murine sequences, they still may elicit an unwanted
`immune reaction, the human anti-chimeric antibody (HACA) reaction, especially when administered for prolonged peri-
`ods, e.g., forchronic indications, such as rheumatoid arthritis (see e.g., Elliott, M.J., et al. (1994) Lancet 344:1 125-1 127;
`Elliot, M.J., et al. (1994) Lancet 344:1105-1 110).
`[0004]
`A preferred hTNFoc inhibitory agent to murine mAbs or derivatives thereof (e.g., chimeric or humanized anti-
`bodies) would be an entirely human anti—hTN For antibody, since such an agent should not elicitthe HAMA reaction, even
`if used for prolonged periods. Human monoclonal autoantibodies against hTNFa have been prepared using human
`hybridoma techniques (Boyle, P., et al. (1993) Cell. Immunol. 152:556-568; Boyle, P., et al. (1993) Cell. Immunol. 152:
`569—581; European Patent Application Publication No. 614 984 A2 by Boyle, et al.). However, these hybridoma—derived
`monoclonal autoantibodies were reported to have an affinity for hTNFoc that was too low to calculate by conventional
`methods, were unable to bind soluble hTNFoc and were unable to neutralize hTNFoc-induced cytotoxicity (see Boyle, et
`al.; supra). Moreover, the success of the human hybridoma technique depends upon the natural presence in human
`peripheral blood of lymphocytes producing autoantibodies specific for hTNFoc. Certain studies have detected serum
`autoantibodies against hTN For in human subjects (Fomsgaard, A., etal. (1 989) Scand. J. Immunol. 30:21 9—223; Bendtzen,
`K., et al. (1990) Prog. Leukocyte Biol. 103:447-452), whereas others have not (Leusch, H-G., et al. (1991) J. Immunol.
`Methods 139:145-147).
`[0005] Alternative to naturally—occurring human anti—hTNFoc antibodies would be a recombinant hTNFoc antibody.
`Recombinant human antibodies that bind hTNFa with relatively low affinity (i.e., Kd ~10'7M) and afast off rate (i.e., Kofi~
`10'2 sec-1) have been described (Griffiths, A.D., et al. (1993) EM BO J. 12:725-734). However, because of their relatively
`fast dissociation kinetics, these antibodies may not be suitable fortherapeutic use. Additionally, a recombinant human
`anti-hTNFa has been described that does not neutralize hTNFoc activity, but rather enhances binding of hTNFoc to the
`surface of cells and enhances internalization of hTNFoc (Lidbury, A., et al. (1994) Biotechnol. Ther. 5:27-45; PCT Pub-
`lication No. WO 92/03145 by Aston, R. et al.)
`[0006] Recombinant human antibodies that bind soluble hTNFa with high affinity and slow dissociation kinetics and
`
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`

`EP 1 528 933 B1
`
`that have the capacity to neutralize hTN For activity, including hTN For-induced cytotoxicity (in vitro and in vivo) and hTN Foc-
`induced cell activation, have also been described (see U.S. Patent No. 6,090,382).
`
`Summary of the Invention
`
`[0007] The scope of the present invention is defined by the claims and any information that does not fall within the
`claims is provided for information only There is a need for a stable aqueous pharmaceutical formulation with an extended
`shelf life, comprising an antibody which is suitable for therapeutic use to inhibit or counteract detrimental hTN For activity.
`There is also a need fora stable aqueous pharmaceutical formulation with an extended shelf life, comprising an antibody
`suitable fortherapeutic use which is easily administered and contains a high protein concentration.
`[0008]
`In particular, the invention provides a liquid aqueous pharmaceutical formulation having a pH of 4 to 8 and
`comprlsmg
`
`(a) 20 to 130 mg/ml of a human IgG 1 anti—Tumor Necrosis Factor alpha (TNFor) antibody comprising a light chain
`variable region comprising the amino acid sequence of SEQ ID NO: 1 and a heavy chain variable region comprising
`the amino acid sequence of SEQ ID NO: 2,
`(b) 10-14 mg/ml of mannitol,
`(c) 0.1 -5 mg/ml of polysorbate 80,
`(cl) 1—1.5 mg/ml of citric acid monohydrate,
`(e) 0.25-0.5 mg/ml of sodium citrate,
`(f) 1.25-1.75 mg/ml ofdisodium phosphate dihydrate,
`(g) 0.7-1.1 mg/ml of sodium dihydrogen phosphate dihydrate, and
`(h) 6.0-6.4 mg/ml sodium chloride.
`
`[0009] This invention also provides a liquid aqueous pharmaceutical formulation consisting of atherapeutically effective
`amount of an antibody in a buffered solution as defined in the claims and having a shelf life of at least 18 months. The
`invention also includes an aqueous pharmaceutical formulation comprising a therapeutically effective amount of an
`antibody in a buffered solution as defined in the claims and having a shelf life of at least 18 months in the liquid state.
`In afurther embodiment, the formulation ofthe invention is stable following at Ieast3 freeze/thaw cycles ofthe formulation.
`In still another embodiment, the antibody is D2E7.
`[0010] This invention also provides a liquid aqueous pharmaceutical formulation comprising atherapeutically effective
`amount of an antibody in a buffered solution as defined in the claims and having enhanced stability of at least 12 months
`at a temperature of2 - 8°C. In one embodiment, the formulation has enhanced stability of at least 1 8 months. In a further
`embodiment, the antibody is D2E7.
`[0011] The invention further provides a liquid aqueous pharmaceutical formulation comprising a therapeutically effec-
`tive amount of an antibody in a buffered solution as defined in the claims which is easily administratable. In a further
`embodiment, the antibody is D2E7.
`[0012]
`In one embodiment of the invention, the liquid aqueous pharmaceutical formulation is suitable for injection. In
`a further embodiment, the formulation is suitable for single use sc injection. In yet another embodiment, the concentration
`of the antibody in the formulation is about 50 mg/ml.
`In still another embodiment, the formulation has a high protein
`concentration. In yet another embodiment of the invention, the formulation is not light sensitive.
`[0013]
`In one embodiment of the invention, the liquid aqueous pharmaceutical formulation as defined in the claims
`contains an antibody, that dissociates from human TNFrx with a Kd of 1 X 10'8 M or less and a Kofl rate constant of 1 x
`10'3 5'1 or less, both determined by surface plasmon resonance, and neutralizes human TN For cytotoxicity in a standard
`in vitro L929 assay with an IC50 of 1 x 10'7 M or less. In another embodiment, the formulation of the invention contains
`an antibody, which dissociates from human TN For with a Kofi rate constant of 5 x104 s1 or less. In a further embodiment,
`the formulation contains an antibody, which dissociates from human TN For with a Koff rate constant of 1 x 10'4 3'1 or
`less. In still afurther embodiment, theformulation of the invention contains an antibody, which neutralizes human TNFoc
`cytotoxicity in astandard in vitro L929 assay with an IC50 of 1 x10'8 M or less. In yet another embodiment ofthe invention,
`the claimed formulation includes an antibody, which neutralizes human TNFa cytotoxicity in a standard in vitro L929
`assay with an IC50 of 1 x10’9 M or less. Another embodiment ofthe invention, includes aformulation where the antibody,
`neutralizes human TNFoc cytotoxicity in a standard in vitro L929 assay with an IC50 of 1 x 10'10 M or less.
`[0014]
`In another embodiment of the invention, the liquid aqueous pharmaceutical formulation contains an antibody,
`which is a recombinant antibody. In another embodiment, the formulation contains an antibody, which inhibits human
`TNFa—induced expression of ELAM-1 on human umbilical vein endothelial cells. In still another embodiment, the claimed
`formulation includes the D2E7 antibody.
`[0015] The antibody as contained in the claimed formulation, has a light chain CDRS domain having the amino acid
`sequence of SEQ ID NO: 3,and has a heavy chain CDR3 domain having the amino acid sequence of SEQ ID NO: 4. In
`
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`

`EP 1 528 933 B1
`
`still another embodiment, the antibodyof the formulation neutralizes the activity of human TN For, chimpanzee TN For and
`at least one additional primate TNFoc selected from the group consisting of baboon TNFoc, mannosetTNFoc, cynomolgus
`TNFoc and rhesus TNFoc.
`In a further embodiment, the formulation of the invention includes an antibody, which also
`neutralizes the activity of mouse TN FOL. The formulation of the invention also an antibody, which neutralizes the activity
`of pig TNFa.
`In one embodiment, the pH of the formulation is between about 4.5 to about 6.0. In another embodiment, the pH is
`between about 4.8 to about 5.5. In yet another embodiment, the pH of the invention is between about 5.0 to about 5.2.
`[0016]
`In one embodiment of the invention, the liquid aqueous pharmaceutical formulation includes about 1.305 mg/ml
`of citric acid, about 0.305 mg/ml of sodium citrate, about 1.53 mg/ml of disodium phosphate dihydrate, about 0.86 mg/ml
`of sodium dihydrogen phosphate dihydrate, and about 6.165 mg/ml of sodium chloride. In yet another embodiment, the
`formulation of the invention includes the antibody D2E7. In yet a further embodiment, the formulation of the invention
`may be administered to a subject suffering from a disorder in which TN Foe activity is detrimental such that TN For activity
`in the subject is inhibited
`
`Detailed Description of the Invention
`
`[0017] The scope of the present invention is defined by the claims and any information that does not fall within the
`claims is provided for information only This invention pertains to a liquid aqueous pharmaceutical formulation as defined
`in the claims with a pH of about 4 to about 8 which contains a high protein concentration, including an antibody, and has
`enhanced stability. This invention also pertains to a liquid aqueous pharmaceutical formulation for therapeutic use in a
`subject suffering from a condition characterized by detrimental TN Fa activity. The formulation of the invention comprises
`the following constituents: an antibody which binds to human TNFoc with high affinity, a low off rate and high neutralizing
`capacity; a buffer, which includes citric acid, sodium citrate, disodium phosphate dihydrate, and sodium dihydrogen
`phosphate dihydrate; tonicity agents, which include mannitol and sodium chloride; a detergent, including polysorbate
`80; and sodium hydroxide, for pH adjustment.
`
`Definitions
`
`In order that the present invention may be more readily understood, certain terms are first defined.
`[0018]
`[0019] The term "subject" is intended to include living organisms, e.g., prokaryotes and eukaryotes. Examples of
`subjects include mammals, e.g., humans, dogs, cows, horses, pigs, sheep, goats, cats, mice, rabbits, rats, andtransgenic
`non—human animals. In specific embodiments of the invention, the subject is a human.
`[0020] The term "pharmaceutical formulation" refers to preparations which are in such form as to permit the biological
`activity of the active ingredients to be unequivocally effective, and which contain no additional components which are
`significantly toxic to the subjects to which the formulation would be administered. "Pharmaceutically acceptable" excip-
`ients (vehicles, additives) are those which can reasonably be administered to a subject mammal to provide an effective
`dose of the active ingredient employed.
`[0021]
`A "stable" formulation is one in which the antibodytherein essentially retains its physical stability and/or chemical
`stability and/or biological activity upon storage. Various analytical techniques for measuring protein stability are available
`in the art and are reviewed in Peptide and Protein Drug Delivery, 247—301, Vincent Lee Ed., Marcel Dekker, Inc., New
`York, N.Y., Pubs. (1 991) and Jones, A. Adv. Drug Delivery Rev. 10: 29-90 (1 993), for example. Stability can be measured
`at a selected temperature for a selected time period. Preferably, the formulation is stable at room temperature (about
`30°C) or at 40°C forat least 1 month and/orstable at about 28°C. for at least 1 year for at least 2 years. Furthermore,
`the formulation is preferably stable following freezing (to, 6.9., -70°C) and thawing of theformulation, hereinafter referred
`to as a "freeze/thaw cycle."
`[0022] An antibody "retains its physical stability" in a pharmaceutical formulation if it shows substantially no signs of
`aggregation, precipitation and/or denaturation upon visual examination of color and/or clarity, or as measured by UV
`light scattering or by size exclusion chromatography.
`[0023] An antibody "retains its chemical stability" in a pharmaceutical formulation, if the chemical stability at a given
`time is such that the antibody is considered to still retain its biological activity as defined below. Chemical stability can
`be assessed by detecting and quantifying chemically altered forms ofthe antibody. Chemical alteration may involve size
`modification (e.g. clipping) which can be evaluated using size exclusion chromatography, SDS-PAGE and/or matrix-
`assisted laser desorption ionization/time—of flight mass spectrometry (MALDl/TOF MS), for example. Other types of
`chemical alteration include charge alteration (e.g. occurring as a result of deamidation) which can be evaluated by ion-
`exchange chromatography, for example.
`[0024] An antibody "retains its biological activity" in a pharmaceutical formulation, if the antibody in a pharmaceutical
`formulation is biologically active for its intended purpose. For example, biological activity is retained if the biological
`activity of the antibody in the pharmaceutical formulation is within about 30%, about 20%, or about 10% (within the errors
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`EP 1 528 933 B1
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`of the assay) of the biological activity exhibited at the time the pharmaceutical formulation was prepared (6.9., as
`determined in an antigen binding assay).
`[0025]
`"Isotonic" is a term recognized in the art. Isotonic can mean, for example, that the formulation of interest has
`essentially the same osmotic pressure as human blood. Isotonic formulations will generally have an osmotic pressure
`from about 250 to 350 mOsm. lsotonicity can be measured using a vapor pressure or ice-freezing type osmometer, for
`example. A "tonicity agent" is a compound which renders the formulation isotonic.
`[0026]
`A "polyol" is asubstance with multiple hydroxyl groups, and includes sugars (reducing and nonreducing sugars),
`sugar alcohols and sugar acids. Preferred polyols herein have a molecular weight which is less than about 600 kD (9.9.
`in the range from about 120 to about 400 kD). A "reducing sugar" is one which contains a hemiacetal group that can
`reduce metal ions or react covalently with lysine and other amino groups in proteins and a "nonreducing sugar" is one
`which does not have these properties of a reducing sugar. Examples of reducing sugars arefructose, mannose, maltose,
`lactose, arabinose, xylose, ribose, rhamnose, galactose and glucose. Nonreducing sugars include sucrose, trehalose,
`sorbose, melezitose and raffinose. Mannitol, xylitol, erythritol, threitol, sorbitol and glycerol are examples of sugar alco-
`hols. As to sugar acids, these include L—gluconate and metallic salts thereof. Where it desired that the formulation is
`freeze-thaw stable, the polyol is preferably one which does not crystallize at freezing temperatures (e.g. -20°C) such
`that it destabilizes the antibody in the formulation. The polyl may also act as a tonicity agent.
`In the invention, one
`ingredient of the formulation is mannitol in a concentration of 10-14 mg/ml.
`[0027] As used herein, "buffer" refers to a buffered solution that resists changes in pH by the action of its acid-base
`conjugate components. The buffer of this invention has a pH in the range from 4 to 8; preferably from about 4.5 to about
`7; and most preferably has a pH in the range from about 5.0 to about 6.5.
`[0028]
`In a pharmacological sense, in the context of the present invention, a "therapeutically effective amount" or
`"effective amount" of an antibody refers to an amount effective in the prevention or treatment of a disorder for the
`treatment of which the antibody is effective. A "disorder" is any condition that would benefit from treatment with the
`antibody. This includes chronic and acute disorders or diseases includingthose pathological conditions which predisposes
`the subject to the disorder in question.
`[0029]
`A "preservative" is a compound which can be included in the formulation to essentially reduce bacterial action
`therein, thus facilitating the production of a multi-use formulation, for example. Examples of potential preservatives
`include octadecyldimethylbenzyl ammonium chloride, hexamethonium chloride, benzalkonium chloride (a mixture of
`alkylbenzyldimethylammonium chlorides in which the alkyl groups are long-chain compounds), and benzethonium chlo-
`ride. Other types of preservatives include aromatic alcohols such as phenol, butyl and benzyl alcohol, alkyl parabens
`such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, 3-pentanol, and m-cresol.
`[0030]
`"Treatment" refers to both therapeutic treatment and prophylactic or preventative measures. Those in need of
`treatment include those already with the disorder as well as those in which the disorder is to be prevented.
`[0031] The phrases "parenteral administration" and "administered parenterally" as used herein means modes of ad-
`ministration otherthan enteral and topical administration, usually by injection, and includes, without limitation, intravenous,
`intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal,
`subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal and intrasternal injection and infusion.
`[0032] The phrases "systemic administration," "administered systemically,
`peripheral administration" and "adminis-
`tered peripherally" as used herein mean the administration of a compound, drug or other material other than directly
`into the central nervous system, such that it enters the patient‘s system and, thus, is subject to metabolism and other
`like processes, for example, subcutaneous administration.
`[0033] The phrase "pharmaceutically acceptable carrier" is art recognized and includes a pharmaceutically acceptable
`material, composition orvehicle, suitable for administration to mammals. The carriers include liquid or solid filler, diluent,
`excipient, solvent or encapsulating material, involved in carrying or transporting the subject agent from one organ, or
`portion of the body, to another organ, or portion of the body. Each carrier must be "acceptable" in the sense of being
`compatible with the other ingredients of the formulation and not injurious to the patient.
`[0034] The term "human TNFa" (abbreviated herein as hTNFa, or simply hTNF), as used herein, is intended to refer
`to a human cytokine that exists as a 17 kD secreted form and a 26 kD membrane associated form, the biologically active
`form of which is composed of a trimer of noncovalently bound 17 kD molecules. The structure of hTNFoz is described
`further in, for example, Pennica, D., et al. (1984) Nature 312:724-729; Davis, J.M., et al. (1987) Biochemistry 26:
`1322—1326; and Jones, E.Y., etal. (1989) Nature 338:225—228. The term human TN For is intended to include recombinant
`human TN For (rhTN Fa), which can be prepared by standard recombinantexpression methods or purchased commercially
`(R & D Systems, Catalog No. 210-TA, Minneapolis, MN).
`[0035] The term "antibody", as used herein, is intendedto referto immunoglobulin molecules comprised offourpolypep-
`tide chains, two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain is
`comprised of a heavy chain variable region (abbreviated herein as HCVR or VH) and a heavy chain constant region.
`The heavy chain constant region is comprised of three domains, CH1, CH2 and CH3. Each light chain is comprised of
`a light chain variable region (abbreviated herein as LCVR or VL) and a light chain constant region. The light chain
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`constant region is comprised of one domain, CL. The VH and VL regions can be further subdivided into regions of
`hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved,
`termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-
`terminus to carboxy-terminus in thefollowing order: FR1, CDR1, FR2, CDR2, FR3, CDRS, FR4. The formulation of the
`disclosure contains an antibody with CDR1, CDR2, and CDR3 sequences like those described in US. Patent Nos.
`6,090,382 and 6,258,562.
`[0036] The term "antigen-binding portion" of an antibody (or simply "antibody portion"), as used herein, refers to one
`or morefragments of an antibodythat retain the abilityto specifically bind to an antigen (e.g., hTN Fa). It has been shown
`that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of
`binding fragments encompassed within the term "antigen-binding portion" of an antibody include (i) a Fab fragment, a
`monovalent fragment consisting of the VL, VH, CL and CH1 domains; (ii) a F(ab')2 fragment, a bivalent fragment com-
`prising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and
`CH1 domains; (iv) a varagment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment
`(Ward et