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`(19)United States
`
`
`(12)Patent Application Publication
`US 2005/0148497 Al
`(10)Pub. No.:
`( 43)Pub. Date:Jul. 7, 2005
`Khan
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`US 20050148497 Al
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`(54) METHOD FOR ADMINISTERING GLP-1
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`
`MOLECULES
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`Related U.S. Application Data
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`(76) Inventor: Mohammed Amin Khan, Carmel, IN
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`filed on Feb.(60) Provisional application No. 60/358,184,
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`20, 2002.
`(US)
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`Publication Classification
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`Correspondence Address:
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`ELI LILLY AND COMPANY
`PATENT DIVISION
`P.O. BOX 6288
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`
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`INDIANAPOLIS, IN 46206-6288 (US)
`
`(21) Appl. No.:
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`1 0/504,717
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`Feb. 7, 2003
`(22) PCT Filed:
`
`(86) PCT No.:
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`PCT/US03/03111
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`.......................... A61K 38/26; A61K 38/17 (51) Int. Cl.7
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`
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`
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`.................................................................. 51 4/8 (52) U.S. Cl.
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`(57)
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`ABSTRACT
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`The invention encompasses formulations that demonstrate
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`the feasibility of oral absorption comprising GLP-1 com­
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`pounds and specified delivery agents.
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` PFIZER, INC. v. NOVO NORDISK A/S - IPR2020-01252, Ex. 1018, p. 1 of 47
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`MOLECULES
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`FIELD OF THE INVENTION
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`BACKGROUND OF THE INVENTION
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`SUMMARY OF THE INVENTION
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`US 2005/0148497 Al
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`Jul. 7, 2005
`
`1
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`
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`METHOD FOR ADMINISTERING GLP-1
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`short in vivo half-life when administered as a solution
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`formulation, suggested that these compounds could not be
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`effectively delivered through the oral route. Thus, it was
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`surprising that GLP-1 compounds could be formulated such
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`that biologically active molecules were absorbed into the
`[0001] The present invention relates to a formulation
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`blood stream after oral administration.
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`useful for the oral administration comprising a glucagon-like
`[0007] The present invention involves the use of specific
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`peptide-1 ( GLP-1) compound and a specified delivery agent.
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`delivery agent molecules that interact with GLP-1 com­
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`Oral administration of the formulations can be used to treat
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`pounds in a non-covalent fashion to allow the compounds to
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`type 2 diabetes as well as a variety of other conditions.
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`cross gut membranes and yet remain therapeutically active.
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`Although the delivery agents employed in the present inven­
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`tion have been disclosed in a series of U.S. patents (see U.S.
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`Pat. Nos. 5,541,155; 5,693,338; 5,976,569; 5,643,957;
`[0002] Over the past several decades, continuous strides
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`5,955,503; 6,100,298; 5,650,386; 5,866,536; 5,965,121;
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`have been made to improve the treatment of diabetes mel­
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`5,989,539; 6,001,347; 6,071,510; 5,820,881; and 6,242,495;
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`litus. Approximately 90% of people with diabetes have type
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`see also WO 02/02509; WO 01/51454; WO 01/44199; WO
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`2 diabetes, also known as non-insulin dependent diabetes
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`01/32130; WO 00/59863; WO 00/50386; WO 00/47188;
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`mellitus (NIDDM). Type 2 diabetics generally still make
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`and WO 00/40203), oral administration of formulations
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`insulin, but the insulin cannot be used effectively by the
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`comprising GLP-1 compounds with these delivery agents
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`body's cells. This is primarily because the amount of insulin
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`has not been disclosed or suggested. Further, numerous
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`produced in response to rising blood sugar levels is not
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`parameters impact whether a particular class of compounds
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`sufficient to allow cells to efficiently take up glucose and
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`can be effectively delivered in combination with one or more
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`thus, reduce blood sugar levels.
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`classes of delivery agents. For example, the conformation of
`[0003] A large body of pre-clinical and clinical research
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`the peptide, the surface charges on the molecule under
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`data suggests that glucagon-like peptide-1 (GLP-1) com­
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`certain formulation conditions, the solubility profile, the
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`pounds show great promise as a treatment for type 2 diabetes
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`stability as a formulated component, as well as susceptibility
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`and other conditions. GLP-1 induces numerous biological
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`to protease digestion and in vivo stability all influence the
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`effects such as stimulating insulin secretion, inhibiting glu­
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`ability to deliver a compound orally.
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`cagon secretion, inhibiting gastric emptying, enhancing glu­
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`cose utilization, and inducing weight loss. Further, pre­
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`clinical studies suggest that GLP-1 may also act to prevent
`[0008] The present invention encompasses the develop­
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`the � cell deterioration that occurs as the disease progresses.
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`ment of novel formulations comprising GLP-1 compounds
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`Perhaps the most salient characteristic of GLP-1 is its ability
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`and delivery agents that can be administered orally. The
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`to stimulate insulin secretion without the associated risk of
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`present invention provides a formulation which can be
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`hypoglycemia that is seen when using insulin therapy or
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`administered orally comprising a GLP-1 compound and a
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`some types of oral therapies that act by increasing insulin
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`specified delivery agent. The GLP-1 compound can be
`expression.
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`native GLP-1; GLP-1 fragments; GLP-1 analogs; GLP-1
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`[0004] However, development of a GLP-1 therapeutic has
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`derivatives of native, fragments, or analogs of GLP-1; and
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`been extremely difficult. This is primarily due to the insta­
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`Exendin-3 and Exendin-4. The delivery agent is selected
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`bility of the peptide during manufacturing processes, in
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`from delivery agents described in U.S. Pat. Nos. 5,541,155;
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`solution formulations, and in vivo. The only published
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`5,693,338; 5,976,569; 5,643,957; 5,955,503; 6,100,298;
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`clinical studies employing GLP-1 compounds to treat hyper­
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`5,650,386; 5,866,536; 5,965,121; 5,989,539, 6,001,347;
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`glycemia or other conditions involve formulating GLP-1
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`6,071,510; 5,820,881; and 6,242,495; and WO 02/02509;
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`compounds such that they can be delivered by subcutaneous
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`WO 01/51454; WO0l/44199; WO0l/32130; WO00/59863;
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`injection or through continuous subcutaneous infusion or
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`WO00/50386; WO00/47188; and WO 00/40203.
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`continuous intravenous administration. Many type 2 diabet­
`[0009] Preferred GLP-1 compounds are analogs or deriva­
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`ics or obese patients desiring to lose weight will not be
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`tives of analogs having modifications at one or more of the
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`willing to undertake a treatment regimen that may involve
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`following positions: 8, 12, 16, 18, 19, 20, 22, 25, 27, 30, 33,
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`several injections per day. Thus, there is a need to develop
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`and 37 and show increased potency compared with Val8-
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`GLP-1 compound therapeutics that can be delivered by an
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`GLP-1(7-37) OH. Preferred GLP-1 compounds are also
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`alternative non-invasive means such as by oral delivery.
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`described in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3,
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`[0005] Unfortunately, there are numerous barriers to effec­
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`SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7,
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`tive oral delivery of peptides. The high acid content and
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`SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID
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`ubiquitous digestive enzymes of the digestive tract will often
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`NO:11, SEQ ID NO:12, SEQ ID NO:13, or SEQ ID NO:14.
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`degrade proteins and peptides before they reach the site of
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`More preferred GLP-1 compounds are described in com­
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`absorption. Further, many peptides cannot effectively
`pounds of SEQ ID NO:2, SEQ ID NO:12, SEQ ID NO:13,
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`traverse the cells of the epithelial membrane in the small
`and SEQ ID NO:14.
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`intestine to reach the bloodstream. Finally, many drugs
`[0010] Preferred delivery agents are described in Table 1.
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`become insoluble at the low pH levels encountered in the
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`More preferred delivery agents are delivery agents corre­
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`digestive tract and, thus, are not absorbed effectively.
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`sponding to numbers of Table 1 selected from the group
`[0006] The fact that GLP-1 compounds are relatively
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`consisting of 1, 2, 4, 5, 6, 9, 10, 11, 13, 14, 15, 20, 21, 22,
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`unstable in solution formulations, only remain in solution
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`23, 24,26, 28,30, 31,35, 36,38, 39,40, 41, 42,43,44,46,
`51, 52, and 54.
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`under a fairly narrow set of conditions, and have a relatively
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`US 2005/0148497 Al
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`Jul. 7, 2005
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`2
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`DETAILED DESCRIPTION OF IBE
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`INVENTION
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`[0011] The present invention also encompasses a method
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`analog or derivative thereof. The nomenclature used to
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`of stimulating the GLP-1 receptor in a subject in need of
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`describe GLP-1 (7-37)OH is also applicable to GLP-1
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`such stimulation, said method comprising the step of admin­
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`fragments. For example, GLP-1(9-36)OH denotes a GLP-1
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`istering to the subject an effective amount of the oral
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`fragment obtained by truncating two amino acids from the
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`formulation described herein. Subjects in need of GLP-1
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`N-terminus and one amino acid from the C-terminus. The
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`receptor stimulation include those with non-insulin depen­
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`amino acids in the fragment are denoted by the same number
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`dent diabetes and obesity.
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`as the corresponding amino acid in GLP-1(7-37)OH. For
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`example, the N-terminal glutamic acid in GLP-1(9-36)OH is
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`at position 9; position 12 is occupied by phenylalanine; and
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`position 22 is occupied by glycine, as in GLP-1(7-37)OH.
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`For GLP-1(7-36)OH, the glycine at position 37 of GLP-1(7-
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`[0012] The three-letter abbreviation code for amino acids
`37)OH is deleted.
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`used in this specification conforms with the list contained in
`[0020] A "GLP-1 analog" has sufficient homology to
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`Table 3 of Annex C, Appendix 2 of the PCT Administrative
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`GLP-1(7-37)OH or a fragment of GLP-1(7-37)OH such that
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`Instructions and with 37 C.F.R. § 1.822(d)(1)(2000).
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`the analog has insulinotropic activity. Preferably, a GLP-1
`[0013] For purposes of the present invention as disclosed
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`analog has the amino acid sequence of GLP-1(7-37)OH or
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`and described herein, the following terms and abbreviations
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`a fragment thereof, modified so that from one, two, three,
`are defined as follows.
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`four or five amino acids differ from the amino acid in
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`corresponding position of GLP-1(7-37)OH or a fragment of
`[0014] The term "formulation" as used herein refers to a
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`GLP-1(7-37)OH. In the nomenclature used herein to desig­
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`GLP-1 compound and a specified delivery agent combined
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`nate GLP-1 compounds, the substituting amino acid and its
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`together which can be administered orally such that GLP-1
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`position is indicated prior to the parent structure. For
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`compound passes through the gut into the systemic circu­
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`example, Glu22-GLP-1(7-37)OH designates a GLP-1 com­
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`lation and has the ability to bind to the GLP-1 receptor and
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`pound in which the glycine normally found at position 22 of
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`initiate a signal transduction pathway resulting in insulino­
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`GLP-1(7-37)OH has been replaced with glutamic acid;
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`tropic activity. The formulation can optionally comprise
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`Val8-Glu22-GLP-1(7-37)OH designates a GLP-1 compound
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`other agents so long as the GLP-1 retains the ability to bind
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`in which alanine normally found at position 8 and glycine
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`the GLP-1 receptor.
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`normally found at position 22 of GLP-1(7-37)OH have been
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`[0015] The term "oral" as used herein refers to delivery of
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`replaced with valine and glutamic acid, respectively.
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`a compound by mouth such that the compound passes
`[0021] GLP-1 molecules also include polypeptides in
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`through the stomach, small intestine, or large intestine into
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`which one or more amino acids have been added to the
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`the systemic circulation.
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`N-terminus and/or C-terminus of GLP-1(7-37)OH, or frag­
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`[0016] The term "GLP-1 compound" as used herein refers
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`ments or analogs thereof. It is preferred that GLP-1 mol­
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`to polypeptides that include naturally occurring GLP-1
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`ecules of this type have up to about thirty-nine amino acids.
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`polypeptides (GLP-1(7-37)OH and GLP-1(7-36)NH2),
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`The amino acids in the "extended" GLP-1 molecule are
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`GLP-1 fragments, GLP-1 analogs, GLP-1 derivatives of
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`denoted by the same number as the corresponding amino
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`naturally occurring GLP-1 polypeptides, GLP-1 fragments,
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`acid in GLP-1(7-37)OH. For example, for a GLP-1 molecule
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`or GLP-1 analogs, and Exendin-3 and Exendin-4 that have
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`obtained by adding two amino acids to the N-terminus of
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`the ability to bind to the GLP-1 receptor and initiate a signal
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`GLP-1(7-37)OH, the N-terminal amino acid is located at
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`transduction pathway resulting in insulinotropic activity.
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`position 5; and for a GLP-1 molecule obtained by adding
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`one amino acid to the C-terminus of GLP-1(7-37)OH, the
`[0017] The term "insulinotropic activity" refers to the
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`C-terminal amino acid is located at position 38. Thus,
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`ability to stimulate insulin secretion in response to elevated
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`position 12 is occupied by phenylalanine and position 22 is
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`glucose levels, thereby causing glucose uptake by cells and
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`occupied by glycine in both of these "extended" GLP-1
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`decreased plasma glucose levels. For example, insulinotro­
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`compounds, as in GLP-1(7-37)OH. Amino acids 1-6 of an
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`pic activity can be determined using the method described in
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`extended GLP-1 molecule are preferably the same as or a
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`Example 1. A GLP-1 molecule has insulinotropic activity if
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`conservative substitution of the amino acid at the corre­
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`islet cells secrete insulin levels in the presence of the GLP-1
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`sponding position of GLP-1(1-37)OH. Amino acids 38-45 of
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`molecule above background levels.
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`an extended GLP-1 molecule are preferably the same as or
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`[0018] The term "DPP IV resistant" refers to GLP-1
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`a conservative substitution of the amino acid at the corre­
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`molecules that have extended metabolic stability and
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`sponding position of glucagon or Exendin-4.
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`improved biological activity. For example, DPP IV resis­
`[0022] A "GLP-1 derivative" refers to a molecule having
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`tance can be determined using the method described in
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`the amino acid sequence of GLP-1, a GLP-1 fragment, or a
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`Example 2. A GLP-1 molecule is DPP Iv resistant if in the
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`GLP-1 analog, but additionally having chemical modifica­
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`presence of DPP IV the GLP-1 molecule has extended
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`tion of one or more of its amino acid side groups, a-carbon
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`metabolic stability above that of native GLP-1. DPP IV
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`atoms, terminal amino group, or terminal carboxylic acid
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`resistant GLP-1 molecules can have an amino acid change at
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`group. A chemical modification includes, but is not limited
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`the DPP IV recognition site (position 8), or DPP IV resistant
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`to, adding chemical moieties, creating new bonds, and
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`peptides can have an attached group that restricts the acces­
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`removing chemical moieties. Modifications at amino acid
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`sibility of the DPP IV to the recognition site, or both.
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`side groups include, without limitation, acylation of lysine
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`[0019] A "GLP-1 fragment" is a polypeptide obtained
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`E-amino groups, N-alkylation of arginine, histidine, or
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`after truncation of one or more amino acids from the
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`lysine, alkylation of glutamic or aspartic carboxylic acid
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`N-terminus and/or C-terminus of GLP-1(7-37)OH or an
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`groups, and deamidation of glutamine or asparagine. Modi-
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`US 2005/0148497 Al
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`Jul. 7, 2005
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`3
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`fications of the terminal amino group include, without
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`by a peptide bond. Peptides can vary in length from dipep­
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`limitation, the des-amino, N-lower alkyl, N-di-lower alkyl,
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`tides with two amino acids to polypeptides with several
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`and N-acyl modifications. Modifications of the terminal
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`hundred amino acids. Preferred peptides include di-peptides,
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`carboxy group include, without limitation, the amide, lower
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`tri-peptides, tetra-peptides, and penta-peptides.
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`alkyl amide, dialkyl amide, and lower alkyl ester modifica­
`[0026] Furthermore, the delivery agents of the present
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`one or more tions. Lower allyl is C1-C4 alkyl. Furthermore,
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`invention are optionally in a salt form. Examples of salts
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`side groups, or terminal groups, may be protected by pro­
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`include sodium, hydrochloric acid, sulfuric acid, phosphoric
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`tective groups known to the ordinarily-skilled protein chem­
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`acid, citric acid, acetic acid, sulfate, phosphate, chloride,
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`ist. The a-carbon of an amino acid may be mono-or
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`bromide, iodide, acetate, propionate, hydrobromic acid,
`dimethylated.
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`sodium hydroxide, potassium hydroxide, ammomum
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`[0023] For the purposes of the present invention, an in
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`hydroxide, and potassium carbonate.
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`vitro GLP-1 receptor-signaling assay is used to determine
`[0027] The various oral formulations of the present inven­
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`whether a particular extended GLP-1 peptide will exhibit
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`tion may optionally encompass a pharmaceutically accept­
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`insulinotropic activity in vivo. Extended GLP-1 peptides
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`able buffer. Examples of pharmaceutically acceptable buff­
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`encompassed by the present invention have an in vitro
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`ers include phosphate buffers such as dibasic sodium
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`potency that is not less than one-tenth the in vitro potency of
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`phosphate, TRIS, glycylglycine, maleate, sodium acetate,
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`the DPP IV resistant GLP-1 analog known as Val8-GLP-
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`sodium citrate, sodium tartrate, or an amino acid such as
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`1(7-37)OH. More preferably, the extended GLP-1 peptides
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`glycine, histidine, lysine or arginine. Other pharmaceutically
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`of the present invention are as potent or more potent than
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`acceptable buffers are known in the art. Preferably, the buffer
`Val8-GLP-1 (7-37)OH.
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`is selected from the group consisting of phosphate, TRIS,
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`maleate, and glycine. Even more preferably the buffer is
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`[0024] "In vitro potency" as used herein is the measure of
`TRIS.
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`the ability of a peptide to activate the GLP-1 receptor in a
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`cell-based assay. In vitro potency is expressed as the "EC50"
`[0028] Preferably, the TRIS concentration is between
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`which is the effective concentration of compound that results
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`about 1 mM and 100 mM. Even more preferably, the
`
`
`in 50% activity in a single dose-response experiment. For
`
`
`concentration is between about 10 mM and about 50 mM,
`
`
`
`
`
`the purposes of the present invention, in vitro potency is
`
`most preferably the buffer is about 20 mM.
`
`
`
`determined using a fluorescence assay that employs HEK-
`[0029] The pH of the oral formulations is adjusted to
`
`
`
`
`293 Aurora CRE-BLAM cells that stably express the human
`
`
`provide stability and to be acceptable for oral administra­
`
`
`
`GLP-1 receptor. These HEK-293 cells have stably integrated
`
`
`
`tion. Preferably, the pH is adjusted to between about 7.0 and
`
`
`
`
`a DNA vector having a cAMP response element (CRE)
`
`
`
`
`about 9.0, more preferably the pH is between about 7.4 and
`
`
`
`
`driving expression of the 3-lactamase (ELAM) gene. The
`
`
`8.4. Even more preferably the pH is between about 7.8 and
`
`
`
`
`interaction of a GLP-1 agonist with the receptor initiates a
`
`
`
`8.4. Most preferably, the pH is between about 7.8 and 8.1.
`
`
`signal that results in activation of the cAMP response
`
`
`
`
`element and subsequent expression of �-lactamase. The
`[0030] The various oral formulations of the present inven­
`
`
`
`
`
`�-lactamase CCF2/AM substrate that emits fluorescence
`
`
`
`
`tion may optionally encompass a suspending agent. Some
`
`
`
`when it is cleaved by �-lactamase (Aurora Biosciences
`
`
`
`delivery agents require a suspending agent due to their
`
`
`
`
`Corp.) can then be added to cells that have been exposed to
`
`
`
`
`solubility characteristics. An example of a suspending agent
`
`
`
`a specific amount of GLP-1 agonist to provide a measure of
`
`
`is hydroxypropylmethylcellulose. Preferably, the final con­
`
`
`
`
`GLP-1 agonist potency. The assay is further described in
`
`
`centration of hydroxypropylmethylcellulose is between
`
`
`Zlokarnik, et al. (1998) Science 279: 84-88 (See also
`
`about 2% and about 10% (weight/volume). Even more
`
`
`
`
`
`
`listed in for the compounds Example 1). The EC50 values
`
`
`
`preferably, the concentration is between about 2% and about
`
`example 1 were determined using the ELAM assay
`
`
`
`5% (w/v). Most preferably the concentration is about 3.9%
`
`
`
`described above by generating a dose response curve using
`(w/v).
`
`
`
`
`dilutions ranging from 0.00003 nanomolar to 30 nanomolar.
`
`
`
`
`Relative in vitro potency values are established by running
`[0031] The oral formulations of the present invention may
`
`
`
`
`
`
`
`Val8-GLP-1(7-37)OH as a control and assigning the control
`
`
`
`optionally comprise a cosolvent. Some delivery agents
`
`a reference value of 1.
`
`
`
`
`require cosolvents due to their solubility characteristics.
`
`
`
`
`Examples of cosolvents include ethanol, N-methylpyrroli­
`
`
`
`[0025] The term "delivery agent" refers to molecules in
`
`done, N,N-dimethylacetamide, N,N-dimethylformamide,
`
`
`
`
`U.S. Pat. Nos. 5,541,155; 5,693,338; 5,976,569; 5,643,957;
`
`
`
`
`
`glycofurol, ethoxydiol, propylene glycol, polyethylene gly­
`
`
`
`5,955,503; 6,100,298; 5,650,386; 5,866,536; 5,965,121;
`
`col 300 and polyvinylpyrrolidone. Preferably, the final con­
`
`
`
`5,989,539; 6,001,347; 6,071,510; 5,820,881; and 6,242,495;
`
`
`
`
`centration of the cosolvents is between about 5% and about
`
`
`
`and WO 02/02509; WO 01/51454; WO 01/44199; WO
`
`
`30% (volume/volume). Even more preferably, the concen­
`
`
`
`01/32130; WO 00/59863; WO 00/50386; WO 00/47188;
`
`
`
`
`tration is between about 10% and about 25% (v/v). Most
`
`
`and WO 00/40203. The delivery agents are generally
`
`
`preferably the concentration is about 20% (v/v).
`
`
`
`derived from amino acids and are useful in the oral formu­
`
`
`lations of the present invention., The derived amino acids
`[0032] The oral formulations of the present invention may
`
`
`
`
`
`
`
`can also be in the form of poly amino acids, and peptides. An
`
`
`
`
`optionally comprise a preservative. Preservative refers to a
`
`
`amino acid is any carboxylic acid having at least one free
`
`
`
`
`compound that is added to the formulation to act as an
`
`
`amine group and includes naturally occurring and synthetic
`
`
`
`
`antimicrobial agent. Among preservatives known in the art
`
`
`
`amino acids. Poly amino acids are either peptides or two or
`
`
`
`as being effective and acceptable in parenteral formulations
`
`
`more amino acids linked by a bond formed by other groups
`
`
`
`are phenolic preservatives, alkylparabens, benzyl alcohol,
`
`
`
`which can be linked, e.g., an ester, anhydride, or an anhy­
`
`
`
`chlorobutanol, resorcinol, and other similar preservatives,
`
`
`dride linkage. Peptides are two or more amino acids joined
`
`
`
`
`
`and various mixtures thereof. Examples of phenolic deriva-
`
` PFIZER, INC. v. NOVO NORDISK A/S - IPR2020-01252, Ex. 1018, p. 4 of 47
`
`

`

`US 2005/0148497 Al
`4
`
`Jul. 7, 2005
`
`[0039] The two naturally occurring truncated GLP-1 pep­
`
`
`
`
`
`
`
`
`tives include cresols and phenol or a mixture of cresols and
`
`
`tides are represented in Formula I, SEQ ID NO:1.
`
`
`
`
`
`phenol. Examples of cresols include meta-cresol, ortho­
`
`
`
`
`cresol, para-cresol, chlorocresol, or mixtures thereof. Alky­
`
`
`
`or mixtures lparaben refers to a C1 to C4 alkylparaben,
`
`
`
`thereof. Examples of alkylparabens include methylparaben,
`
`
`
`ethylparaben, propylparaben, or butylparaben. The concen­
`
`
`
`
`
`trations must be sufficient to maintain preservative effec­
`
`
`
`
`tiveness by retarding microbial growth. Preferably, the pre­
`
`
`
`
`servative is a phenol derivative. More preferably the
`
`
`
`
`preservative is a cresol. Even more preferably the preserva­
`tive is meta-cresol.
`
`His7-Ala-Glu-Gly10-Thr-Phe-Thr-Ser-Asp-Val-Ser­
`
`
`
`Ser-Tyr-Leu20-Glu-Gly-Gln-Ala-Ala25-Lys-Glu-Phe­
`
`Ile-Ala30-Trp-Leu-Val-Lys-Gly35-Arg-Xaa37
`
`FORMULA I
`
`SEQ ID NO: 1
`
`FORMULA I
`
`(SEQ ID NO: 2)
`
`
`
`
`
`[0040] wherein:
`[0033] A preferred concentration of a preservative in the
`
`
`
`final mixture is about 1.0 mg/mL to about 20.0 mg/mL.
`
`[0041] Xaa37 is Gly, or -NH2.
`
`
`
`More preferred ranges of concentration of preservative in
`[0042] Preferably, a GLP-1 compound has the amino acid
`
`
`
`the final mixture are about 2.0 mg/mL to about 8.0 mg/mL,
`
`
`
`
`sequence of SEQ ID NO: 1 or is modified so that from one,
`about 2.5 mg/mL to about 4.5 mg/mL and about 2.0 mg/mL
`
`
`two, three, four or five amino acids differ from SEQ ID NO:
`
`to about 4.0 mg/mL. A most preferred concentration of
`1.
`
`
`
`preservative in the final mixture is about 3.0 mg/mL.
`[0043] A preferred group of GLP-1 compounds is com­
`
`
`
`
`posed of GLP-1 analogs of Formula I (SEQ ID NO:2).
`
`
`
`[0034] The oral formulations of the present invention may
`
`
`
`
`optionally comprise an isotonicity agent. Isotonicity agents
`
`
`
`refer to compounds that are tolerated physiologically and
`
`
`
`
`impart a suitable tonicity to the formulation to prevent the
`
`
`
`net flow of water across cell membranes. Examples of such
`
`
`
`
`compounds include glycerin, salts, e.g., NaCl, and sugars,
`
`
`
`
`
`e.g., dextrose, mannitol, and sucrose. These compounds are
`1 7
`20
`21
`22
`23
`24
`18
`19
`-
`Xaa
`-Xaa
`-Xaa
`-Xaa
`-Xaa
`-Xaa
`-Xaa
`-Xaa
`
`
`
`commonly used for such purposes at known concentrations.
`32
`27
`25
`26
`30
`31
`33
`
`
`
`One or more isotonicity agents may be added to adjust the
`
`
`
`
`
`
`
`Xaa-Xaa -Xaa -Phe-Ile-Xaa -xaa-Xaa-Xaa -
`40
`37
`34
`35
`36
`38
`39
`41
`
`
`
`
`ionic strength or tonicity. The preferred isotonicity agent is
`-
`
`
`
`
`
`
`
`Xaa-Xaa -Xaa -Xaa-Xaa-Xaa -Xaa-Xaa
`
`
`
`NaCl. The concentration of the NaCl is preferably between
`
`about 10 mM and 200 mM, more preferred is between about
`
`
`50 mM and 150 mM, and most preferred is about 100 mM.
`
`14
`16-
`
`His-Xaa-Xaa-Gly-Xaa-Phe-Thr-Xaa-Asp-Xaa
`
`8
`
`9
`
`11
`
`Xaa
`
`42
`43
`44
`-Xaa
`-Xaa
`-Xaa
`
`45
`
`[0044] wherein:
`[0035] The administration compositions may alternatively
`
`
`
`
`
`be in the form of a solid, such as a tablet, capsule or particle,
`Asp, or Lys;
`
`
`such as a powder. Solid dosage forms may be prepared by
`
`
`mixing the solid form of the compound with the solid form
`[0046] Xaa9 is Glu, Asp, or Lys;
`
`
`
`of the active agent. Alternatively, a solid may be obtained
`
`
`
`
`
`from a solution of compound and active agent by methods
`Asp, or Lys;
`
`
`
`known in the art, such as freeze drying, precipitation,
`
`crystallization ad solid dispersion.
`
`[0045] Xaa8 is Ala, Gly, Ser, Thr, Leu, Ile, Val, Glu,
`
`Val, Glu, [0047] Xaa11 is Thr, Ala, Gly, Ser, Leu, Ile,
`
`[0048] Xaa14 is Ser, Ala, Gly, Thr, Leu, Ile, Val, Glu,
`
`
`
`[0036] GLP-1 Compounds Appropriate for use m the
`
`Present Invention:
`
`[0050] Xaa17 is Ser, Ala, Gly, Thr, Leu, Ile, Val, Glu,
`
`[0051] Xaa18 is Ser, Ala, Gly, Thr, Leu, Ile, Val, Glu,
`
`[0053] Xaa20 is Leu, Ala, Gly, Ser, Thr, Ile, Val, Glu,
`
`
`
`[0055] Xaa22 is Gly, Ala, Ser, Thr, Leu, Ile, Val, Glu,
`
`
`Asp, or Lys;
`[0049] Xaa16 is Val, Ala, Gly, Ser, Tor, Leu, Ile, Tyr,
`
`Glu, Asp, Trp, or Lys;
`
`
`invention [0037] The GLP-1 compounds of the present
`
`
`
`
`can be made by a variety of methods known in the art such
`Asp, or Lys;
`
`
`
`as solid-phase synthetic chemistry, purification of GLP-1
`
`
`
`
`molecules from natural sources, recombinant DNA technol­
`
`
`
`ogy, or a combination of these methods. For example,
`Asp, Trp, Tyr, or Lys;
`
`
`
`
`methods for preparing GLP-1 peptides are described in U.S.
`[0052] Xaa19 is Tyr, Phe, Trp, Glu, Asp, Gln, or Lys;
`
`
`
`Pat. Nos. 5,118,666; 5,120,712; 5,512,549; 5,977,071; and
`6,191,102.
`
`
`
`Asp, Met, Trp, Tyr, or Lys;
`[0038] By custom in the art, the amino terminus of GLP-
`
`
`[0054] Xaa21 is Glu, Asp,
`or Lys;
`
`
`1(7-37)OH has been assigned number residue 7, and the
`
`
`carboxy-terminus has been assigned number 37. The other
`
`
`
`amino acids in the polypeptide are numbered consecutively,
`Asp, or Lys;
`
`
`
`as shown in SEQ ID NO:l. For example, position 12 is
`[0056] Xaa23 is Gln, Asn, Arg, Glu, Asp, or Lys;
`
`
`phenylalanine and position 22 is glycine.
`
` PFIZER, INC. v. NOVO NORDISK A/S - IPR2020-01252, Ex. 1018, p. 5 of 47
`
`

`

`
`Asp, or Lys;
`
`7
`
`
`
`8
`
`9
`
`11
`
`12
`
`
`
`
`
`
`
`
`
`US 2005/0148497 Al
`
`Jul. 7, 2005
`
`5
`
`[0057] Xaa24 is Ala, Gly, Ser, Thr, Leu, Ile, Val, Arg,
`
`The compounds of formula I derivatized as described in WO
`
`
`
`
`Glu, Asp, or Lys;
`
`
`99/43706 and underivatized are encompassed by the present
`invention.
`
`[0058] Xaa25 is Ala, Gly, Ser, Thr, Leu, Ile, Val, Glu,
`Asp, or Lys;
`[0080] Another preferred group of GLP-1 compounds is
`
`
`
`
`
`
`composed of GLP-1 analogs of formula II (SEQ ID NO:3):
`[0059] Xaa26 is Lys, Arg, Gln, Glu, Asp, or His;
`
`
`[0060] Xaa27 is Leu, Glu, Asp, or Lys;
`
`FORMULA II
`
`[0061] Xaa30 is Ala, Gly, Ser, Thr, Leu, Ile, Val, Glu,
`
`(SEQ ID NO: 3)
`16-
`Xaa -Xaa -Xaa-Gly-Xaa -Xaa--Thr-Ser-Asp-Xaa
`
`[0062] Xaa31 is Trp, Phe, Tyr, Glu, Asp, or Lys;
`
`26-
`Ser-Xaa -Xaa -Leu-Glu-Gly-Xaa-Xaa -Ala-Xaa
`
`18
`
`19
`
`2 3
`
`24
`
`
`
`
`
`
`
`
`
`[0063] Xaa32 is Leu, Gly, Ala, Ser, Thr, Ile, Val, Glu,
`
`
`Asp, or Lys;
`
`27
`
`30
`
`31
`
`
`
`Xaa-Phe-I le-Xaa-xaa -Leu-Xaa -Xaa-Xaa
`
`
`
`
`
`33
`
`34
`
`35
`-
`
`[0064] Xaa33 is Val, Gly, Ala, Ser, Thr, Leu, Ile, Glu,
`
`
`Asp, or Lys;
`
`[0081] wherein:
`[0065] Xaa34 is Asn, Lys, Arg, Glu, Asp, or His;
`
`[0082] Xaa7 is: L-histidine, D-histidine, desamino­
`
`
`
`
`[0066] Xaa35 is Gly, Ala, Ser, Thr, Leu, Ile, Val, Glu,
`
`histidine, 2-amino-histidine, �-hydroxy-histidine,
`Asp, or Lys;
`
`
`homohistidine, a-fluoromethyl-histidine or a-me­
`thyl-histidine;
`
`[0067] Xaa36 is Gly, Arg, Lys, Glu, Asp, or His;
`
`[0068] Xaa37 is Pro, Gly, Ala, Ser, Thr, Leu, Ile, Val,
`
`Glu, Asp, or Lys, or is deleted;
`[0069] Xaa38 is Ser, Arg, Lys, Glu, Asp, or His, or is
`deleted;
`[0070] Xaa39 is Ser, Arg, Lys, Glu, Asp, or His, or is
`deleted;
`
`[0083] Xaa8 is: Gly, Ala, Val, Leu, Ile, Ser, or Thr;
`
`
`[0084] Xaa9 is: Thr, Ser,Arg, Lys, Trp, Phe, Tyr, Glu,
`or His;
`
`[0085] Xaa11 is: Asp, Glu, Arg, Thr, Ala, Lys, or His;
`
`
`[0086] Xaa12 is: His, Trp, Phe, or Tyr;
`[0087] Xaa16 is: Leu, Ser, Thr, Trp, His, Phe, Asp,
`[0071] Xaa40 is Gly, Asp, Glu, or Lys, or is deleted;
`
`Val, Tyr, Glu, or Ala;
`[0088] Xaa18 is: His, Pro, Asp, Glu, Arg, Ser, Ala, or
`Lys;
`[0073] Xaa42 is Ser, Pro, Lys, Glu, or Asp, or 1s
`[0089] Xaa19 is: Gly, Asp, Glu, Gln, Asn, Lys, Arg, or
`Cys;
`[0074] Xaa43 is Ser, Pro, Glu, Asp, or Lys, or 1s
`deleted;
`[0090] Xaa23 is: His, Asp, Lys, Glu, Gln, or Arg;
`
`
`[0072] Xaa41 is Ala, Phe, Trp, Tyr, Glu, Asp, or Lys,
`or is deleted;
`
`
`deleted;
`
`
`
`deleted; and
`
`[0076] Xaa45 is Ala, Ser, Val, Glu, Asp, or Lys,
`Ala-NH2, Ser-NH2, Val-NH2, Glu-NH2,
`
`[0075] Xaa44 is Gly, Pro, Glu, Asp, or Lys, or 1s
`
`[0091] Xaa24 is: Glu, Arg, Ala, or Lys;
`
`[0092] Xaa26 is: Trp, Tyr, Phe, Asp, Lys, Glu, or His;
`[0093] Xaa27 is: Ala, Glu, His, Phe, Tyr, Trp, Arg, or
`Lys;
`
`that [0077] Asp-NH2, or Lys-NH2, or is deleted; provided
`
`
`
`
`when the amino acid at position 37, 38, 39, 40, 41, 42, 43,
`[0094] Xaa30 is: Ala, Glu, Asp, Ser, or His;
`
`or 44 is deleted, then each amino acid downstream of that
`[0095] Xaa31 is: Asp, Glu, Ser, Thr, Arg, Trp, or Lys;
`amino acid is also deleted.
`
`[0078] It is preferred that the GLP-1 compound of formula
`
`
`
`[0096] Xaa33 is: Asp, Arg, Val, Lys, Ala, Gly, or Glu;
`
`I contain less than six amino acids that differ from the
`[0097] Xaa34 is: Glu, Lys, or Asp;
`
`
`corresponding amino acid in GLP-1(7-37)OH or Exendin-4.
`
`
`It is more preferred that less than five amino acids differ
`[0098] Xaa35 is: Thr, Ser, Lys, Arg, Trp, Tyr, Phe,
`
`
`
`from the corresponding amino acid in GLP-1(7-37)OH or
`Asp, Gly, Pro, His, or Glu;
`
`
`
`Exendin-4. It is even more preferred that less than four
`
`amino acids differ from the corresponding amino acid in
`
`GLP-1(7-37)OH or Exendin-4.
`
`
`
`
`Glu, or His;
`
`[0099] Xaa36 is: Tor, Ser, Asp, Tip, Tyr, Phe, Arg,
`
`[0079] GLP-1 compounds of the present invention include
`
`
`
`
`[0100] R37 is: Lys, Arg, Thr, Ser, Glu, Asp, Trp, Tyr,
`
`
`
`
`derivatives of formula I such as a C-1-6-ester, or amide, or
`
`Phe, His, Gly, Gly-Pro, or is deleted.
`
`
`
`C-1-6-alkylamide, or C-1-6-dialkylamide thereof. WO99/
`
`
`
`
`43706 describes derivatives of GLP-1 compounds of for­
`[0101] Another preferred group of GLP-1 compounds is
`
`
`
`
`
`mula I and is incorporated by reference herein in its entirety.
`
`
`
`composed of GLP-1 analogs of Formula III (SEQ ID NO:4):
`
` PFIZER, INC. v. NOVO NORDISK A/S - IPR2020-01252, Ex. 1018, p. 6 of 47
`
`

`

`US 2005/0148497 A1
`
`Jul. 7, 2005
`
`FORMULA III
`
`(SEQ ID NO : 4)
`Xaa-Xaa-Glu-Gly-Xaa-Xaa'?-Thr-Ser-Asp-Xaa
`Ser-Ser-Tyr-Leu-Glu-Xaa’-Xaa-Xaa-Xaa’-Lys
`Xaa-Phe-Ile-Xaa-Trp-Leu-Xaa-Xaa-Xaa
`
`Xaa-R7
`
`0102)
`wherein:
`0103) Xaa" is: L-histidine, D-histidine, desamino
`histidine, 2-amino-histidine, B-hydroxy-histidine,
`homohistidine, C.-fluoromethylhistidine or C.-meth
`ylhistidine;
`0104 Xaa is: Gly, Ala, Val, Leu, Ile, Ser, or Thr;
`y
`0105 Xaa' is: Asp, Gl