WO1998008872A1 - Derives de glp-2 - Google Patents

Derives de glp-2 Download PDF

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Publication number
WO1998008872A1
WO1998008872A1 PCT/DK1997/000360 DK9700360W WO9808872A1 WO 1998008872 A1 WO1998008872 A1 WO 1998008872A1 DK 9700360 W DK9700360 W DK 9700360W WO 9808872 A1 WO9808872 A1 WO 9808872A1
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WO
WIPO (PCT)
Prior art keywords
glp
group
derivative according
lys
lipophilic substituent
Prior art date
Application number
PCT/DK1997/000360
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English (en)
Inventor
Liselotte Bjerre Knudsen
Per Olaf Huusfeldt
Per Franklin Nielsen
Lars Thim
Original Assignee
Novo Nordisk A/S
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Novo Nordisk A/S filed Critical Novo Nordisk A/S
Priority to JP10511193A priority Critical patent/JP2000517308A/ja
Priority to AU41124/97A priority patent/AU4112497A/en
Priority to EP97938802A priority patent/EP0929576A1/fr
Publication of WO1998008872A1 publication Critical patent/WO1998008872A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/26—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/28—Insulins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/04—Anorexiants; Antiobesity agents

Definitions

  • the present invention relates to novel derivatives of human glucagon-like peptide-2 (hGLP-2) and analogues thereof and fragments thereof and analogues of such fragments which have a protracted profile of action and to methods of making and using them.
  • hGLP-2 human glucagon-like peptide-2
  • Peptides are widely used in medical practice, and since they can be produced by recombinant DNA technology it can be expected that their importance will increase also in the years to come. When native peptides or analogues thereof are used in therapy it is generally found that they have a high clearance. A high clearance of a therapeutic agent is inconvenient in cases where it is desired to maintain a high blood level thereof over a prolonged period of time since repeated administrations will then be necessary.
  • peptides which have a high clearance are: ACTH, corticotropin-releasing factor, angiotensin, calcitonin, insulin, glucagon, glucagon-like peptide-1 , glucagon-like peptide-2, insulin-like growth factor-1 , insulin-like growth factor-2, gastric inhibitory peptide, growth hormone- releasing factor, pituitary adenylate cyclase activating peptide, secretin, enterogastrin, somatostatin, somatotropin, somatomedin, parathyroid hormone, thrombopoietin, erythropoietin, hypothalamic releasing factors, prolactin, thyroid stimulating hormones, endorphins, enkephalins, vasopressin, oxytocin, opiods and analogues thereof, superoxide dismutase, interferon, asparaginase, arginase, arginine deamina
  • GLP-2 and other preproglucagon fragments are given i.a. by Schmidt et al. (Diabetologia 28 704-707 (1985). Little is known about the physical chemical properties of GLP-2 but GLP-2 is excpected, like GLP-1, to be a highly flexible and unstable molecule. GLP-2 and fragments thereof and analogues of GLP-2 and fragments thereof are potentially useful i.a. in regulation of appetite and in the treatment of small bowel syndrome. However, the high clearance limits the usefulness of these compounds, and thus there still is a need for improvements in this field.
  • Preproglucagon from which GLP-2 originates, is synthesized i.a. in the L-cells in the distal illeum, in the pancreas and in the brain. Processing of preproglucagon to give GLP-1 and GLP-2 occurs mainly in the L-cells.
  • GLP-2 is a 34 amino acid residue peptide.
  • a simple system is used to describe fragments, analogues and derivatives of GLP-2.
  • Lys 20 GLP-2(1-33) designates a fragment of GLP-2 formally derived from GLP-2 by deleting the amino acid residues No. 34 and substituting the naturally occurring amino acid residue in position 20 (Arg) by Lys.
  • Arg ⁇ Lys ⁇ N'-tetradecanoy GLP-IO-S ⁇ designates a derivative of a GLP-2 analogue formally derived from GLP-2 by C-terminal addition of a Lys residue, exchange of the naturally occurring amino acid residue in position 30 (Lys) with an Arg residue and tetradecanoylation of the ⁇ -amino group of the Lys residue in position 35.
  • the present invention relates to derivatives of GLP-2 and analogues thereof.
  • the derivatives according to the invention have interesting pharmacological properties, in particular they have a more protracted profile of action than the parent peptides.
  • GLP-2 designates human GLP-2.
  • an analogue is used to designate a peptide wherein one or more amino acid residues of the parent peptide have been substituted by another amino acid residue and/or wherein one or more amino acid residues of the parent peptide have been deleted and/or wherein one or more amino acid residues have been added to the parent peptide.
  • derivative is used in the present text to designate a peptide in which one or more of the amino acid residues have been chemically modified, e.g. by alkylation, acylation, ester formation or amide formation.
  • a GLP derivative is used in the present text to designate a derivative of GLP-2 or an analogue thereof.
  • the parent peptide from which such a derivative is formally derived is in some places referred to as the "GLP moiety" of the derivative.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent attached to any one amino acid residue.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent attached to any one amino acid residue with the proviso that only if the substituent has an ⁇ -carboxylic acid group or is an alkyl group can it be attached to the N-terminal or C-terminal amino acid residue of the parent peptide.
  • the present invention relates to a GLP-2 derivative wherein the lipophilic substituent comprises from 4 to 40 carbon atoms, more preferred from 8 to 25 carbon atoms.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to an amino acid residue in such a way that a carboxyl group of the lipophilic substituent forms an amide bond with an amino group of the amino acid residue.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to an amino acid residue in such a way that an amino group of the lipophilic substituent forms an amide bond with a carboxyl group of the amino acid residue.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer which is an unbranched alkane ⁇ , ⁇ -dicarboxylic acid group having from 1 to 7 methylene groups, preferably two methylene groups which spacer forms a bridge between an amino group of the parent peptide and an amino group of the lipophilic substituent.
  • a spacer which is an unbranched alkane ⁇ , ⁇ -dicarboxylic acid group having from 1 to 7 methylene groups, preferably two methylene groups which spacer forms a bridge between an amino group of the parent peptide and an amino group of the lipophilic substituent.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer which is an amino acid residue except Cys, or a dipeptide such as Gly-Lys.
  • a dipeptide such as Gly-Lys is used to designate a dipeptide wherein the C- terminal amino acid residue is Lys, His or Trp, preferably Lys, and wherein the N-terminal amino acid residue is selected from the group comprising Ala, Arg, Asp, Asn, Gly, Glu, Gin, lie, Leu.Val, Phe and Pro.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys and wherein a carboxyl group of the parent peptide forms an amide bond with an amino group of a Lys residue or a dipeptide containing a Lys residue, and the other amino group of the Lys residue or a dipeptide containing a Lys residue forms an amide bond with a carboxyl group of the lipophilic substituent.
  • a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys and wherein a carboxyl group of the parent peptide forms an amide bond with an amino group of a Lys residue or a dipeptide containing a Lys residue, and the other amino group of the Lys residue or a dipeptide containing a Lys residue forms an amide bond with a
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys and wherein an amino group of the parent peptide forms an amide bond with a carboxylic group of the amino acid or dipeptide spacer, and an amino group of the amino acid or dipeptide spacer forms an amide bond with a carboxyl group of the lipophilic substituent.
  • a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys
  • an amino group of the parent peptide forms an amide bond with a carboxylic group of the amino acid or dipeptide spacer
  • an amino group of the amino acid or dipeptide spacer forms an amide bond with a carboxyl group of the lipophilic substituent.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys and wherein a carboxyl group of the parent peptide forms an amide bond with an amino group of the amino acid or dipeptide spacer, and the carboxyl group of the amino acid or dipeptide spacer forms an amide bond with an amino group of the lipophilic substituent.
  • the present invention relates to a GLP-2 derivative wherein a lipophilic substituent is attached to the parent peptide by means of a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys, and wherein a carboxyl group of the parent peptide forms an amide bond with an amino group of Asp or Glu, or a dipeptide containing an Asp or Glu residue, and a carboxyl group of the spacer forms an amide bond with an amino group of the lipophilic substituent.
  • a spacer which is an amino acid residue except Cys, or is a dipeptide such as Gly-Lys
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which comprises a partially or completely hydrogenated cyclopentano- phenathrene skeleton.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a straight-chain or branched alkyl group.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is the acyl group of a straight-chain or branched fatty acid.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is an acyl group selected from the group comprising CH 3 (CH 2 ) n CO-, wherein n is 4 to 38, preferably CH 3 (CH 2 ) 6 CO-, CH 3 (CH 2 ) 8 CO-, CH 3 (CH 2 ) 10 CO-, CH 3 (CH 2 ) 12 CO-, CH 3 (CH 2 ) 14 CO-, CH 3 (CH 2 ) 16 CO-, CH 3 (CH 2 ) 1 ⁇ CO-, CH 3 (CH 2 ) 20 CO- and CH 3 (CH 2 ) 22 CO-.
  • a lipophilic substituent which is an acyl group selected from the group comprising CH 3 (CH 2 ) n CO-, wherein n is 4 to 38, preferably CH 3 (CH 2 ) 6 CO-, CH 3 (CH 2 ) 8 CO-, CH 3 (CH 2 ) 10 CO-, CH 3 (CH 2 ) 12 CO-, CH 3 (CH 2 ) 14 CO
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is an acyl group of a straight-chain or branched alkane ⁇ , ⁇ - dicarboxylic acid.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is an acyl group selected from the group comprising HOOC(CH 2 ) m CO-, wherein m is 4 to 38, preferably HOOC(CH 2 ) 14 CO-, HOOC(CH 2 ) 16 CO-, HOOC(CH 2 ) 18 CO-, HOOCfCH ⁇ CO- and HOOC(CH 2 ) 22 CO-.
  • a lipophilic substituent which is an acyl group selected from the group comprising HOOC(CH 2 ) m CO-, wherein m is 4 to 38, preferably HOOC(CH 2 ) 14 CO-, HOOC(CH 2 ) 16 CO-, HOOC(CH 2 ) 18 CO-, HOOCfCH ⁇ CO- and HOOC(CH 2 ) 22 CO-.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula CH 3 (CH 2 ) p ((CH 2 ) q COOH)CHNH- CO(CH 2 ) 2 CO-, wherein p and q are integers and p+q is an integer of from 8 to 40, preferably from 12 to 35.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula CH 3 (CH 2 ) r CO- NHCH(COOH)(CH 2 ) 2 CO-, wherein r is an integer of from 10 to 24.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula CH 3 (CH 2 ) s CO- NHCH((CH 2 ) 2 COOH)CO-, wherein s is an integer of from 8 to 24.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula COOH(CH 2 ),CO- wherein t is an integer of from 8 to 24.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula -NHCH(COOH)(CH 2 ) 4 NH- CO(CH 2 ) u CH 3 , wherein u is an integer of from 8 to 18.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula -NHCH(COOH)(CH 2 ) 4 NH- COCH((CH 2 ) 2 COOH)NH-CO(CH 2 ) w CH 3 , wherein w is an integer of from 10 to 16.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula -NHCH(COOH)(CH 2 ) 4 NH- CO(CH 2 ) 2 CH(COOH)NH-CO(CH 2 ) !( CH 3 , wherein x is an integer of from 10 to 16.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which is a group of the formula -NHCH(COOH)(CH 2 ) 4 NH- CO(CH 2 ) 2 CH(COOH)NHCO(CH 2 ) y CH 3 , wherein y is zero or an integer of from 1 to 22.
  • the present invention relates to a GLP-2 derivative which has one lipophilic substituent.
  • the present invention relates to a GLP-2 derivative which has two lipophilic substituents.
  • the present invention relates to a GLP-2 derivative in which the C-terminal amino acid residue is present in the form of the amide.
  • the present invention relates to a GLP-2 derivative having a lipophilic substituent which can be negatively charged.
  • the present invention relates to a GLP-2 derivative the parent peptide of which is selected from the group comprising GLP-2(1-35) or an analogue thereof.
  • the present invention relates to a GLP-2 derivative derived from a GLP-2 fragment selected from the group comprising GLP-2(1-30); GLP-2(1- 31 ); GLP-2(1-32); GLP-2(1-33); GLP-2(1-34) and GLP-2(1-35).
  • the present invention relates to a GLP-2 derivative wherein the designation analogue comprises derivatives wherein a total of up to ten amino acid residues have been exchanged with any ⁇ -amino acid residue.
  • the present invention relates to a derivative of a GLP-2 analogue wherein the designation analogue implies that the parent peptide is human GLP-2 wherein a total of up to six, more preferred up to three, amino acid residues have been added, deleted or substituted with other amino acid residues which can be coded for by the genetic code.
  • the present invention relates to a GLP-2 derivative wherein the parent peptide is selected from the group comprising Lys 20 GLP-2(1-33) and Lys 20 Arg 30 GLP-2(1-33).
  • the present invention relates to a GLP-2 derivative wherein the parent peptide is Arg 30 Lys 3 GLP-2(1-34).
  • the present invention relates to a GLP-2 derivative wherein the parent peptide is selected from the group comprising Arg 30 Lys 35 GLP-2(1-35); Arg 30 ' 35 Lys 20 GLP-2(1-35) and Arg 35 GLP-2(1-35).
  • the present invention relates to a GLP-2 derivative which is selected from the group comprising
  • Lys 20 (N ⁇ -tetradecanoyl)GLP-2(1-33);
  • Lys 20 (N c -tetradecanoyl)Arg 30 GLP-2(1-33);
  • Lys 20 (N ⁇ -( ⁇ -carboxynonadecanoyl))GLP-2(1-33);
  • Lys 20 (N e -( ⁇ -carboxynonadecanoyl))Arg 30 GLP-2(1-33); Arg 30 Lys 35 (N ⁇ -( ⁇ -carboxynonadecanoyl))GLP-2(1-35);
  • the present invention relates to a pharmaceutical composition
  • a pharmaceutical composition comprising a GLP-2 derivative and a pharmaceutically acceptable vehicle or carrier.
  • the present invention relates to the use of a GLP-2 derivative according to the invention for the preparation of a medicament which has a more protracted action than the parent peptide. In a further preferred embodiment, the present invention relates to the use of a GLP-2 derivative according to the invention for the preparation of a medicament with protracted effect for the treatment of obesity.
  • the present invention relates to the use of a GLP-2 derivative according to the invention for the preparation of a medicament with protracted effect for the treatment of small bowel syndrome.
  • the lipophilic substituent attached to the GLP-2 moiety preferably comprises 4-40 carbon atoms, in particular 8-25 carbon atoms.
  • the lipophilic substituent may be attached to an amino group of the GLP-2 moiety by means of a carboxyl group of the lipophilic substituent which forms an amide bond with an amino group of the amino acid to which it is attached.
  • the lipophilic substituent may be attached to said amino acid in such a way that an amino group of the lipophilic substituent forms an amide bond with a carboxyl group of the amino acid.
  • the lipophililic substituent may be linked to the GLP-2 moiety via an ester bond.
  • the ester can be formed either by reaction between a carboxyl group of the GLP-2 moiety and a hydroxyl group of the substituent-to-be or by reaction between a hydroxyl group of the GLP-2 moiety and a carboxyl group of the substituent-to-be.
  • the lipophilic substituent can be an alkyl group which is introduced into a primary amino group of the GLP-2 moiety.
  • the lipophilic substituent is attached to the GLP-2 moiety by means of a spacer in such a way that a carboxyl group of the spacer forms an amide bond with an amino group of the GLP-2 moiety.
  • suitable spacers are succinic acid, Lys, Glu or Asp, or a dipeptide such as Gly-Lys.
  • the spacer is succinic acid, one carboxyl group thereof may form an amide bond with an amino group of the amino acid residue, and the other carboxyl group thereof may form an amide bond with an amino group of the lipophilic substituent.
  • the spacer is Lys, Glu or Asp
  • the carboxyl group thereof may form an amide bond with an amino group of the amino acid residue
  • the amino group thereof may form an amide bond with a carboxyl group of the lipophilic substituent.
  • a further spacer may in some instances be inserted between the ⁇ -amino group of Lys and the lipophilic substituent.
  • such a further spacer is succinic acid which forms an amide bond with the ⁇ - amino group of Lys and with an amino group present in the lipophilic substituent.
  • such a further spacer is Glu or Asp which forms an amide bond with the ⁇ -amino group of Lys and another amide bond with a carboxyl group present in the lipophilic substituent, that is, the lipophilic substituent is a N c -acylated lysine residue.
  • the lipophilic substituent has a group which can be negatively charged.
  • One preferred group which can be negatively charged is a carboxylic acid group.
  • the parent peptide can be produced by a method which comprises culturing a host cell containing a DNA sequence encoding the peptide and capable of expressing the peptide in a suitable nutrient medium under conditions permitting the expression of the peptide, after which the resulting peptide is recovered from the culture.
  • the medium used to culture the cells may be any conventional medium suitable for growing the host cells, such as minimal or complex media containing appropriate supplements. Suitable media are available from commercial suppliers or may be prepared according to published recipes (e.g. in catalogues of the American Type Culture Collection).
  • the peptide produced by the cells may then be recovered from the culture medium by conventional procedures including separating the host cells from the medium by centrifugation or filtration, precipitating the proteinaceous components of the supernatant or filtrate by means of a salt, e.g. ammonium sulphate, purification by a variety of chromatographic procedures, e.g. ion exchange chromatography, gelfiltration chromatography, affinity chromatography, or the like, dependent on the type of peptide in question.
  • a salt e.g. ammonium sulphate
  • the DNA sequence encoding the parent peptide may suitably be of genomic or cDNA origin, for instance obtained by preparing a genomic or cDNA library and screening for DNA sequences coding for all or part of the peptide by hybridisation using synthetic oligonucleotide probes in accordance with standard techniques (see, for example, Sambrook, J, Fritsch, EF and Maniatis, T, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, New York, 1989).
  • the DNA sequence encoding the peptide may also be prepared synthetically by established standard methods, e.g.
  • the DNA sequence may also be prepared by polymerase chain reaction using specific primers, for instance as described in US 4,683,202 or Saiki et al., Science 239 (1988), 487 - 491.
  • the DNA sequence may be inserted into any vector which may conveniently be subjected to recombinant DNA procedures, and the choice of vector will often depend on the host cell into which it is to be introduced.
  • the vector may be an autonomously replicating vector, i.e. a vector which exists as an extrachromosomal entity, the replication of which is independent of chromosomal replication, e.g. a plasmid.
  • the vector may be one which, when introduced into a host cell, is integrated into the host cell genome and replicated together with the chromosome(s) into which it has been integrated.
  • the vector is preferably an expression vector in which the DNA sequence encoding the peptide is operably linked to additional segments required for transcription of the DNA, such as a promoter.
  • the promoter may be any DNA sequence which shows transcriptional activity in the host cell of choice and may be derived from genes encoding proteins either homologous or heterologous to the host cell. Examples of suitable promoters for directing the transcription of the DNA encoding the peptide of the invention in a variety of host cells are well known in the art, cf. for instance Sambrook et al., supra.
  • the DNA sequence encoding the peptide may also, if necessary, be operably connected to a suitable terminator, polyadenylation signals, transcriptional enhancer sequences, and translational enhancer sequences.
  • the recombinant vector of the invention may further comprise a DNA sequence enabling the vector to replicate in the host cell in question.
  • the vector may also comprise a selectable marker, e.g. a gene the product of which complements a defect in the host cell or one which confers resistance to a drug, e.g. ampicillin, kanamycin, tetracyclin, chloramphenicol, neomycin, hygromycin or methotrexate.
  • a secretory signal sequence also known as a leader sequence, prepro sequence or pre sequence
  • the secretory signal sequence is joined to the DNA sequence encoding the peptide in the correct reading frame. Secretory signal sequences are commonly positioned 5' to the DNA sequence encoding the peptide.
  • the secretory signal sequence may be that normally associated with the peptide or may be from a gene encoding another secreted protein.
  • the host cell into which the DNA sequence or the recombinant vector is introduced may be any cell which is capable of producing the present peptide and includes bacteria, yeast, fungi and higher eukaryotic cells.
  • suitable host cells well known and used in the art are, without limitation, E. coli, Saccharomyces cerevisiae, or mammalian BHK or CHO cell lines.
  • the GLP-2 derivatives of the invention can be prepared by introducing the lipophilic substituent into the parent GLP-2 or GLP-2 analogue using methods known per se, see for example WO 95/07931, the contents of which is hereby incorporated in its entirety by reference.
  • N E -acylation of a Lys residue can be carried out by using an activated amide of the acyl group to be introduced as the acylating agent, e.g. the amide with benzotriazole.
  • the acylation is carried out in a polar solvent in the presence of a base.
  • compositions containing a GLP-2 derivative according to the present invention may be administered parenterally to patients in need of such a treatment.
  • Parenteral administration may be performed by subcutaneous, intramuscular or intravenous injection by means of a syringe, optionally a pen-like syringe.
  • parenteral administration can be performed by means of an infusion pump.
  • a further option is a composition which may be a powder or a liquid for the administration of the GLP-2 derivative in the form of a nasal or pulmonal spray.
  • the GLP-2 derivatives of the invention can also be administered transdermally, e.g. from a patch, optionally a iontophoretic patch, or transmucosally, e.g. bucally.
  • compositions containing a GLP-2 derivative of the present invention may be prepared by conventional techniques, e.g. as described in Remington's Pharmaceutical Sciences. 1985 or in Remington: The Science and Practice of Pharmacy, 19 th edition, 1995.
  • the injectable compositions of the GLP-2 derivative of the invention can be prepared using the conventional techniques of the pharmaceutical industry which involves dissolving and mixing the ingredients as appropriate to give the desired end product.
  • the GLP-2 derivative is dissolved in an amount of water which is somewhat less than the final volume of the composition to be prepared.
  • An isotonic agent, a preservative and a buffer is added as required and the pH value of the solution is adjusted - if necessary - using an acid, e.g. hydrochloric acid, or a base, e.g. aqueous sodium hydroxide as needed.
  • the volume of the solution is adjusted with water to give the desired concentration of the ingredients.
  • isotonic agents sodium chloride, mannitol and glycerol.
  • preservatives examples include phenol, m-cresol, methyl p-hydroxybenzoate and benzyl alcohol.
  • Suitable buffers are sodium acetate and sodium phosphate.
  • solutions containing a GLP-2 derivative according to the present invention may also contain a surfactant in order to improve the solubility and/or the stability of the derivative.
  • a composition for nasal administration of GLP-2 may, for example, be prepared as described in European Patent No. 272097 (to Novo Nordisk A S) or in WO 93/18785.
  • the GLP-2 derivatives of this invention can be used in the treatment of various diseases.
  • the particular GLP-2 derivative to be used and the optimal dose level for any patient will depend on the disease to be treated and on a variety of factors including the efficacy of the specific peptide derivative employed, the age, body weight, physical activity, and diet of the patient, on a possible combination with other drugs, and on the severity of the case. It is recommended that the dosage of the GLP-2 derivative of this invention be determined for each individual patient by those skilled in the art in a similar way as for known parent peptides.
  • NMP N-Methyl-2-pyrrolidone.
  • EDPA N-Ethyl-N,N-diisopropylamine.
  • TFA Trifluoroacetic acid.
  • Myr-ONSu Tetradecanoic acid 2,5-dioxopyrrolidin-1-yl ester. Abbreviations:
  • PDMS Plasma Desorption Mass Spectrometry
  • HPLC High Performance Liquid Chromatography amu: atomic mass units
  • the reaction was quenched by the addition of a solution of glycine (4.5 mg, 4.5 ⁇ mol) in 50% aqueous ethanol (451 ⁇ l).
  • the reaction mixture was purified by column chromatography using a cyanopropyl column (Zorbax 300SB-CN) and a standard acetonitrile/TFA system. The column was heated to 65°C and the acetonitrile gradient was 0-100% in 60 minutes. The title compound ( 5.0 mg, 47 %) was isolated from the eluate.

Abstract

Ces dérivés de hGLP-2 (peptide 2 humain de type glucagon), des analogues et fragments de ceux-ci, ainsi que des analogues de tels fragments présentant un substituant lipophile, possèdent des propriétés pharmacologiques intéressantes, notamment un profil d'action plus étalé que celui des peptides parents.
PCT/DK1997/000360 1996-08-30 1997-09-01 Derives de glp-2 WO1998008872A1 (fr)

Priority Applications (3)

Application Number Priority Date Filing Date Title
JP10511193A JP2000517308A (ja) 1996-08-30 1997-09-01 Ｇｌｐ―２誘導体
AU41124/97A AU4112497A (en) 1996-08-30 1997-09-01 Glp-2 derivatives
EP97938802A EP0929576A1 (fr) 1996-08-30 1997-09-01 Derives de glp-2

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
DK0931/96 1996-08-30
DK93196 1996-08-30
DK1259/96 1996-11-08
DK125996 1996-11-08

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WO1999043361A1 (fr) * 1998-02-27 1999-09-02 Novo Nordisk A/S Derives de glp-2 avec teneur en structures en helices superieure a 25 %, formant des agregats de type micellaire partiellement structures
WO2000055119A1 (fr) * 1999-03-17 2000-09-21 Novo Nordisk A/S Methode d'acylation de peptides et nouveaux agents d'acylation
WO2002040537A2 (fr) * 2000-11-16 2002-05-23 Novo Nordisk A/S Analogues et derives du peptide liberateur de gastrine (gpr)
WO2002066511A2 (fr) * 2001-02-16 2002-08-29 Conjuchem Inc. Peptide 2 de type glucagon (glp-2) de longue duree utilise dans le traitement des maladies et troubles gastro-intestinaux
US6444788B1 (en) 1999-03-15 2002-09-03 Novo Nordisk A/S Ion exchange chromatography of GLP-1, analogs and derivatives thereof
US6451974B1 (en) 1999-03-17 2002-09-17 Novo Nordisk A/S Method of acylating peptides and novel acylating agents
JP2002539219A (ja) * 1999-03-15 2002-11-19 ノボ ノルディスク アクティーゼルスカブ Ｇｌｐ−１及び近縁ペプチドのイオン交換クロマトグラフィー分離
US6583111B1 (en) 1996-11-05 2003-06-24 Eli Lilly And Company Use of GLP-1 analogs and derivative adminstered peripherally in regulation of obesity
WO2004029077A2 (fr) * 2002-09-25 2004-04-08 Novo Nordisk A/S Procede de production de peptides acyles
WO2004035624A2 (fr) * 2002-10-14 2004-04-29 Novo Nordisk A/S Composes glp-2, leurs formulations et leurs utilisations
US6770620B2 (en) 2000-09-18 2004-08-03 Sanos Bioscience A/S Use of GLP for the treatment, prevention, diagnosis, and prognosis of bone-related and nutrition-related disorders
WO2004085471A2 (fr) * 2003-03-24 2004-10-07 Novo Nordisk A/S Derives de glp-2
WO2004089985A1 (fr) * 2003-04-11 2004-10-21 Novo Nordisk A/S Compositions pharmaceutiques stables
WO2005027978A2 (fr) * 2003-09-19 2005-03-31 Novo Nordisk A/S Nouveaux derives de glp-1
WO2005028516A2 (fr) * 2003-09-19 2005-03-31 Novo Nordisk A/S Nouvelles etiquettes d'affinite pour les proteines plasmiques
WO2005082404A2 (fr) * 2004-02-27 2005-09-09 Novo Nordisk A/S Composes de glp-2, formulations et utilisations de ceux-ci
WO2007084460A2 (fr) 2006-01-18 2007-07-26 Qps, Llc Compositions pharmaceutiques a stabilite amelioree
US7268113B2 (en) 2001-02-02 2007-09-11 Conjuchem Biotechnologies Inc. Long lasting growth hormone releasing factor derivatives
US7273921B2 (en) 2002-09-25 2007-09-25 Novo Nordisk A/S Method for producing acylated peptides
US7276590B1 (en) 1999-03-15 2007-10-02 Novo Nordisk A/S Ion exchange chromatography of proteins and peptides
WO2008025856A2 (fr) 2006-09-01 2008-03-06 Novo Nordisk Health Care Ag Protéines modifiées
US7371721B2 (en) 2000-09-18 2008-05-13 Sanos Bioscience A/S Use of GLP-2 and related compounds for the treatment, prevention, diagnosis, and prognosis of bone-related disorders and calcium homeostasis related syndromes
EP2033662A1 (fr) 2004-01-21 2009-03-11 Novo Nordisk Health Care AG Conjugaison au moyen de transglutaminase de peptides
US7572884B2 (en) 2001-07-24 2009-08-11 Novo Nordisk A/S Method for making acylated polypeptides
US7595172B2 (en) 2001-07-24 2009-09-29 Novo Nordisk A/S Method for making acylated polypeptides
US7749955B2 (en) 2003-08-21 2010-07-06 Novo Nordisk A/S Separation of polypeptides comprising a racemized amino acid
US7785297B2 (en) 2003-02-04 2010-08-31 Novo Nordisk A/S Injection device with rotatable dose setting
US8158583B2 (en) 2003-11-13 2012-04-17 Novo Nordisk A/S Soluble pharmaceutical compositions for parenteral administration comprising a GLP-1 peptide and an insulin peptide of short time action for treatment of diabetes and bulimia
US8206735B2 (en) 2006-07-11 2012-06-26 Foresee Pharmaceuticals, Llc Pharmaceutical compositions for sustained release delivery of peptides
US8513192B2 (en) 2009-01-22 2013-08-20 Novo Nordisk A/S Stable growth hormone compounds resistant to proteolytic degradation
US8536122B2 (en) 2005-03-18 2013-09-17 Novo Nordisk A/S Acylated GLP-1 compounds
US8603972B2 (en) 2005-03-18 2013-12-10 Novo Nordisk A/S Extended GLP-1 compounds
US8637647B2 (en) 2008-09-12 2014-01-28 Novo Nordisk A/S Method of acylating a peptide or protein
US8779109B2 (en) 2010-01-22 2014-07-15 Novo Nordisk Health Care Ag Growth hormones with prolonged in-vivo efficacy
US8841249B2 (en) 2009-08-06 2014-09-23 Novo Nordisk A/S Growth hormones with prolonged in-vivo efficacy
US8865868B2 (en) 2008-08-06 2014-10-21 Novo Nordisk Healthcare Ag Conjugated proteins with prolonged in vivo efficacy
US9211342B2 (en) 2010-01-22 2015-12-15 Novo Nordisk Healthcare Ag Stable growth hormone compounds resistant to proteolytic degradation
US11045523B2 (en) 2013-04-05 2021-06-29 Novo Nordisk Healthcare Ag Formulation of growth hormone albumin-binder conjugate

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WO2005046716A1 (fr) * 2003-11-13 2005-05-26 Novo Nordisk A/S Compositions pharmaceutiques solubles destinées à l'administration parentérale comprenant un peptide glp-1 et un peptide d'insuline à temps d'action de courte durée, utiles pour le traitement du diabète et de la boulimie

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US5512549A (en) * 1994-10-18 1996-04-30 Eli Lilly And Company Glucagon-like insulinotropic peptide analogs, compositions, and methods of use
WO1996032414A1 (fr) * 1995-04-14 1996-10-17 1149336 Ontario Inc. Peptide-2 analogue au glucagon et ses utilisations therapeutiques

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WO1991011457A1 (fr) * 1990-01-24 1991-08-08 Buckley Douglas I Analogues de glp-1 utiles dans le traitement du diabete
US5512549A (en) * 1994-10-18 1996-04-30 Eli Lilly And Company Glucagon-like insulinotropic peptide analogs, compositions, and methods of use
WO1996032414A1 (fr) * 1995-04-14 1996-10-17 1149336 Ontario Inc. Peptide-2 analogue au glucagon et ses utilisations therapeutiques

Cited By (66)

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Publication number Priority date Publication date Assignee Title
US7211557B2 (en) 1996-11-05 2007-05-01 Eli Lilly And Company Use of GLP-1 analogs and derivatives administered peripherally in regulation of obesity
US6583111B1 (en) 1996-11-05 2003-06-24 Eli Lilly And Company Use of GLP-1 analogs and derivative adminstered peripherally in regulation of obesity
WO1999043361A1 (fr) * 1998-02-27 1999-09-02 Novo Nordisk A/S Derives de glp-2 avec teneur en structures en helices superieure a 25 %, formant des agregats de type micellaire partiellement structures
US7276590B1 (en) 1999-03-15 2007-10-02 Novo Nordisk A/S Ion exchange chromatography of proteins and peptides
US8067554B2 (en) 1999-03-15 2011-11-29 Novo Nordisk A/S Ion exchange chromatography of GLP-1, analogs and derivatives thereof
JP2002539219A (ja) * 1999-03-15 2002-11-19 ノボ ノルディスク アクティーゼルスカブ Ｇｌｐ−１及び近縁ペプチドのイオン交換クロマトグラフィー分離
JP4732590B2 (ja) * 1999-03-15 2011-07-27 ノボ ノルディスク アクティーゼルスカブ Ｇｌｐ−１及び近縁ペプチドのイオン交換クロマトグラフィー分離
EP2348044A1 (fr) 1999-03-15 2011-07-27 Novo Nordisk A/S Chromatographie d'échange d'ions de GLP-1, analogues et dérivés associés
US6444788B1 (en) 1999-03-15 2002-09-03 Novo Nordisk A/S Ion exchange chromatography of GLP-1, analogs and derivatives thereof
EP1956000A1 (fr) 1999-03-17 2008-08-13 Novo Nordisk A/S Agents d'acylation utilisés dans l'acylation de peptides
US6451974B1 (en) 1999-03-17 2002-09-17 Novo Nordisk A/S Method of acylating peptides and novel acylating agents
WO2000055119A1 (fr) * 1999-03-17 2000-09-21 Novo Nordisk A/S Methode d'acylation de peptides et nouveaux agents d'acylation
US6770620B2 (en) 2000-09-18 2004-08-03 Sanos Bioscience A/S Use of GLP for the treatment, prevention, diagnosis, and prognosis of bone-related and nutrition-related disorders
US7371721B2 (en) 2000-09-18 2008-05-13 Sanos Bioscience A/S Use of GLP-2 and related compounds for the treatment, prevention, diagnosis, and prognosis of bone-related disorders and calcium homeostasis related syndromes
WO2002040537A2 (fr) * 2000-11-16 2002-05-23 Novo Nordisk A/S Analogues et derives du peptide liberateur de gastrine (gpr)
WO2002040537A3 (fr) * 2000-11-16 2002-09-26 Novo Nordisk As Analogues et derives du peptide liberateur de gastrine (gpr)
US7268113B2 (en) 2001-02-02 2007-09-11 Conjuchem Biotechnologies Inc. Long lasting growth hormone releasing factor derivatives
US7112567B2 (en) 2001-02-16 2006-09-26 Conjuchem Inc. Long lasting glucagon-like peptide 2 (glp-2) for the treatment of gastrointestinal diseases and disorders
WO2002066511A2 (fr) * 2001-02-16 2002-08-29 Conjuchem Inc. Peptide 2 de type glucagon (glp-2) de longue duree utilise dans le traitement des maladies et troubles gastro-intestinaux
EP1980572A1 (fr) * 2001-02-16 2008-10-15 ConjuChem Biotechnologies Inc. Peptide 2 (GLP-2) de type glucagon longue durée pour le traitement de maladies et de troubles gastrointestinaux
US7737251B2 (en) 2001-02-16 2010-06-15 Conjuchem Biotechnologies Inc. Long lasting glucagon-like peptide 2 (GLP-2) for the treatment of gastrointestinal diseases and disorders
WO2002066511A3 (fr) * 2001-02-16 2003-03-06 Conjuchem Inc Peptide 2 de type glucagon (glp-2) de longue duree utilise dans le traitement des maladies et troubles gastro-intestinaux
US8835132B2 (en) 2001-07-24 2014-09-16 Novo Nordisk A/S Method for making acylated polypeptides
US7595172B2 (en) 2001-07-24 2009-09-29 Novo Nordisk A/S Method for making acylated polypeptides
US7572884B2 (en) 2001-07-24 2009-08-11 Novo Nordisk A/S Method for making acylated polypeptides
US7273921B2 (en) 2002-09-25 2007-09-25 Novo Nordisk A/S Method for producing acylated peptides
US9562910B2 (en) 2002-09-25 2017-02-07 Novo Nordisk A/S Method for producing acylated peptides
WO2004029077A3 (fr) * 2002-09-25 2004-05-13 Novo Nordisk As Procede de production de peptides acyles
WO2004029077A2 (fr) * 2002-09-25 2004-04-08 Novo Nordisk A/S Procede de production de peptides acyles
WO2004035624A2 (fr) * 2002-10-14 2004-04-29 Novo Nordisk A/S Composes glp-2, leurs formulations et leurs utilisations
WO2004035624A3 (fr) * 2002-10-14 2004-09-10 Novo Nordisk As Composes glp-2, leurs formulations et leurs utilisations
US7411039B2 (en) 2002-10-14 2008-08-12 Novo Nordisk A/S GLP-2 compounds, formulations, and uses thereof
US8419692B2 (en) 2003-02-04 2013-04-16 Novo Nordisk A/S Injection device with rotatable dose setting
US7785297B2 (en) 2003-02-04 2010-08-31 Novo Nordisk A/S Injection device with rotatable dose setting
WO2004085471A3 (fr) * 2003-03-24 2004-11-04 Novo Nordisk As Derives de glp-2
WO2004085471A2 (fr) * 2003-03-24 2004-10-07 Novo Nordisk A/S Derives de glp-2
US7595293B2 (en) 2003-04-11 2009-09-29 Novo Nordisk A/S Stable pharmaceutical compositions
WO2004089985A1 (fr) * 2003-04-11 2004-10-21 Novo Nordisk A/S Compositions pharmaceutiques stables
US7749955B2 (en) 2003-08-21 2010-07-06 Novo Nordisk A/S Separation of polypeptides comprising a racemized amino acid
WO2005028516A2 (fr) * 2003-09-19 2005-03-31 Novo Nordisk A/S Nouvelles etiquettes d'affinite pour les proteines plasmiques
EP2932981A3 (fr) * 2003-09-19 2016-02-24 Novo Nordisk A/S Nouveaux dérivés GLP-1 se liant à l'albumine
JP2007505840A (ja) * 2003-09-19 2007-03-15 ノボ　ノルディスク　アクティーゼルスカブ 新規ｇｌｐ−１誘導体
AU2004273573B2 (en) * 2003-09-19 2010-04-22 Novo Nordisk A/S Albumin-binding derivatives of therapeutic peptides
WO2005028516A3 (fr) * 2003-09-19 2005-09-29 Novo Nordisk As Nouvelles etiquettes d'affinite pour les proteines plasmiques
US7897560B2 (en) 2003-09-19 2011-03-01 Novo Nordisk A/S Plasma protein affinity tags
WO2005027978A2 (fr) * 2003-09-19 2005-03-31 Novo Nordisk A/S Nouveaux derives de glp-1
WO2005027978A3 (fr) * 2003-09-19 2005-05-19 Novo Nordisk As Nouveaux derives de glp-1
US8158583B2 (en) 2003-11-13 2012-04-17 Novo Nordisk A/S Soluble pharmaceutical compositions for parenteral administration comprising a GLP-1 peptide and an insulin peptide of short time action for treatment of diabetes and bulimia
EP2368579A1 (fr) 2004-01-21 2011-09-28 Novo Nordisk Health Care AG Conjugaison au moyen de transglutaminase de peptides
EP2033662A1 (fr) 2004-01-21 2009-03-11 Novo Nordisk Health Care AG Conjugaison au moyen de transglutaminase de peptides
WO2005082404A3 (fr) * 2004-02-27 2005-12-01 Novo Nordisk As Composes de glp-2, formulations et utilisations de ceux-ci
WO2005082404A2 (fr) * 2004-02-27 2005-09-09 Novo Nordisk A/S Composes de glp-2, formulations et utilisations de ceux-ci
US8603972B2 (en) 2005-03-18 2013-12-10 Novo Nordisk A/S Extended GLP-1 compounds
US8536122B2 (en) 2005-03-18 2013-09-17 Novo Nordisk A/S Acylated GLP-1 compounds
WO2007084460A2 (fr) 2006-01-18 2007-07-26 Qps, Llc Compositions pharmaceutiques a stabilite amelioree
US8206735B2 (en) 2006-07-11 2012-06-26 Foresee Pharmaceuticals, Llc Pharmaceutical compositions for sustained release delivery of peptides
US8840915B2 (en) 2006-07-11 2014-09-23 Foresec Pharmaceuticals, LLC Pharmaceutical compositions for sustained release delivery of peptides
WO2008025856A2 (fr) 2006-09-01 2008-03-06 Novo Nordisk Health Care Ag Protéines modifiées
US8865868B2 (en) 2008-08-06 2014-10-21 Novo Nordisk Healthcare Ag Conjugated proteins with prolonged in vivo efficacy
US8637647B2 (en) 2008-09-12 2014-01-28 Novo Nordisk A/S Method of acylating a peptide or protein
US8791236B2 (en) 2008-09-12 2014-07-29 Novo Nordisk A/S Method of acylating a peptide or protein
US8513192B2 (en) 2009-01-22 2013-08-20 Novo Nordisk A/S Stable growth hormone compounds resistant to proteolytic degradation
US8841249B2 (en) 2009-08-06 2014-09-23 Novo Nordisk A/S Growth hormones with prolonged in-vivo efficacy
US9211342B2 (en) 2010-01-22 2015-12-15 Novo Nordisk Healthcare Ag Stable growth hormone compounds resistant to proteolytic degradation
US8779109B2 (en) 2010-01-22 2014-07-15 Novo Nordisk Health Care Ag Growth hormones with prolonged in-vivo efficacy
US11045523B2 (en) 2013-04-05 2021-06-29 Novo Nordisk Healthcare Ag Formulation of growth hormone albumin-binder conjugate

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EP0929576A1 (fr) 1999-07-21
JP2000517308A (ja) 2000-12-26
AU4112497A (en) 1998-03-19

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