WO1997046252A1 - Growth hormone component and bone anti-resorptive agent in cyclic (coherence) treatment of osteoporosis - Google Patents

Growth hormone component and bone anti-resorptive agent in cyclic (coherence) treatment of osteoporosis Download PDF

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Publication number
WO1997046252A1
WO1997046252A1 PCT/DK1997/000239 DK9700239W WO9746252A1 WO 1997046252 A1 WO1997046252 A1 WO 1997046252A1 DK 9700239 W DK9700239 W DK 9700239W WO 9746252 A1 WO9746252 A1 WO 9746252A1
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Prior art keywords
growth hormone
phe
2nal
osteoporosis
lys
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PCT/DK1997/000239
Other languages
French (fr)
Inventor
Erik Fink Eriksen
Anne-Marie Kappelgaard
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Novo Nordisk A/S
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Publication date
Application filed by Novo Nordisk A/S filed Critical Novo Nordisk A/S
Priority to EP97924934A priority Critical patent/EP0907374A1/en
Priority to JP10500108A priority patent/JP2000512274A/en
Priority to CA002257174A priority patent/CA2257174A1/en
Priority to IL12698397A priority patent/IL126983A0/en
Priority to AU30259/97A priority patent/AU3025997A/en
Priority to BR9709499A priority patent/BR9709499A/en
Publication of WO1997046252A1 publication Critical patent/WO1997046252A1/en
Priority to NO985573A priority patent/NO985573L/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/02—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing at least one abnormal peptide link
    • C07K5/0207—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing at least one abnormal peptide link containing the structure -NH-(X)4-C(=0), e.g. 'isosters', replacing two amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/16—Amides, e.g. hydroxamic acids
    • A61K31/165—Amides, e.g. hydroxamic acids having aromatic rings, e.g. colchicine, atenolol, progabide
    • 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
    • A61K38/27—Growth hormone [GH] (Somatotropin)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
    • A61P19/10—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease for osteoporosis

Definitions

  • the present invention relates to the treatment or prevention of osteoporosis
  • BSU Bone Multicellular Unit
  • osteoblasts have receptors for both growth hormone and insulin growth factors I and II (IGF-I and IGF-II) (B ⁇ xen et al , Potential Use of Growth Hormone in the Treatment of Osteoporosis, in Abstracts, Workshop on Growth No 5, September 25-26, 1992, Moltkes Palace, Copenhagen, Denmark) Although growth hormone has been found to increase bone mass considerably in rats and dogs, it does not seem to significantly affect bone mass in humans
  • the object of the present invention is to provide a more efficient method of treating or preventing the development of osteoporosis in humans which may ensure a more efficient use of estrogen in order to increase the bone formation over the level obtainable hitherto
  • the present invention relates, in a first aspect, to the treatment of osteoporosis in an animal and to the use of a growth hormone or a compound having growth hormone like or growth hormone releasing effect as well as somatostatin antagonists (in the following also collectively referred to in a generic way as the growth hormone component) and a composition with anti- resorptive action on bone (in the following referred to in a generic way as the estrogenic component) in the treatment
  • the invention relates to increasing the bone mass and preventing the development of osteoporosis in an animal and to the use of a growth hormone component and an estrogenic component in the treatment
  • the present invention relates to a method of treating as well as a method of preventing the development of osteoporosis and related disorders such as osteopenia in an animal compri ⁇ sing simultaneous administration to the animal of amounts of a growth hormone component and an estrogen component effective, in combination, to increase the bone mass in the animal
  • animal used in the present context includes, but is not limited to birds, such as chicken, ducks or turkeys, fish such as salmon, trout or tuna, and mammals such as cows, horses, sheep and human beings
  • the animal to be treated is preferably a mammal and more preferred a human being, most preferably a woman, and the growth hormone is preferably human growth hormone
  • the growth hormone component is administered in a cyclic way which means that periods in which growth hormone component is administered alternate with periods where no growth hormone component is administered as further specified in the present text
  • the estrogen component is preferably administered daily
  • each medicament may, individually, be administered by injection, in the form of a patch for topical application, by nasal administration or in the form of a tablet
  • growth hormone is preferably administered by injection It could also be envisaged that growth hormone was used in a sustained release formulation
  • the estrogen is given continuously during the period of treatment whereas the growth hormone is given at intervals
  • a preferred method of administration comprises daily administration of estrogen and penodical administration of growth hormone for a period of from about 2 days to about 28 days, preferably from about 2 days to about 14 days, more preferred from about 3 days to about 10 days, in particular from about 3 days to about 7 days, with intervals of from about 1 week to about 26 weeks, preferably from about 3 weeks to about 26 weeks, more preferably from about 6 to about 12 weeks, even more preferred from about 6 weeks to about 10 weeks, in particular about 8 weeks, between the periods in which of growth hormone is administered
  • the estrogen component is administered in the amount from 0.001 mg to 10 mg/kg body weight/day When the estrogen component used is estradiol, the total amount given per day is from about 0.5 mg to about 4 mg, preferably from about 1 mg to about 2 mg.
  • estrogen is considered to encompass any preparation containing an estrogenic substance such as a natural human estrogen such as estrone, 17- ⁇ -estrad ⁇ ol and estradiol or derivatives thereof being cleaved in vivo to form natural estrogens, natural equine estrogens prepared from urine from horses, or artificial estrogens without steroid structure such as dienestrol
  • the "estrogen” could also be selected from the group of non-steroidal estrogen directed therapeutics (NSERTs) such as Centchroman, Levormeloxifene, Raloxifene, Droloxifene, Tamoxifene, Idoxifene, or the like or structurally related compounds thereof, such as e g the compounds disclosed in WO 96/21656, WO 95/10513, US Patent 5,280,040, WO 96/09040, EP 0693488, WO 95/34557, EP 0617030, WO 93/10741, US Patent 5,254,568, EP 0683170, EP 06
  • growth hormone may be growth hormone of any origin such as avian, bovine, equine, human, ovine, porcine, salmon, trout or tuna growth hormone, preferably bovine, human or porcine growth hormone, human growth hormone being most preferred
  • the growth hormone used in accordance with the invention may be native growth hormone isolated from a natural source, e g by extracting pituitary glands in a conventional manner, or a growth hormone produced by recombinant techniques, e g as described in E.B Jensen and S Carlsen in Biotech and Bioeng 3S, 1-11 (1990)
  • the growth hormone may also be a truncated form of growth hormone wherein one or more ammo acid residues has (have) been deleted, an analogue thereof wherein one or more ammo acid residues in the native molecule has (have) been substituted by another ammo acid residue, preferably a natural ammo acid residue, as long as the substitution does not have any adverse effect such as antigenicity or essentially
  • GHRH growth hormone releasing hormone
  • growth hormone releasing factor growth hormone releasing factor
  • suitable growth hormone components are growth hormone (GH), IGF-I, IGF-ll, PACAP, GHRH, truncated GHRH, GHRP-1 , GHRP-2, GHRP-6, MK-677 disclosed in R P Margund et al , PNAS 92(15), 1995, p 7001, hexare n disclosed in WO 91/18016, as well as compounds disclosed in WO 95/17422, WO 95/17423, WO 96/05195, WO 96/22997, WO 96/24580, WO 96/24587, WO 97/00894, such compounds are
  • H-H ⁇ s ⁇ (CH 2 NH)D-Trp-Ala-Trp-D-Phe-Lys-NH 2 H-H ⁇ s-D-Trp ⁇ (CH 2 NH)Ala-Trp-D-Phe-Lys-NH 2l H-H ⁇ s-D-Trp-Ala ⁇ (CH 2 NH)Trp-D-Phe-Lys-NH 2 , H-H ⁇ s-D-Trp-Ala-Trp ⁇ (CH 2 NH)D-Phe-Lys-NH 2 , H-His-D-Trp-Ala-Trp-D-Phe ⁇ (CH 2 NH)Lys-NH 2 , H-D-Ala-D-2Nal-Ala ⁇ (CH 2 NH)Trp-D-Phe-Lys-NH 2 , H-D-Ala-D-2Nal-Ala-Trp-D-Phe ⁇ (CH 2 NH
  • P ⁇ pe ⁇ d ⁇ ne-4-carboxyl ⁇ c acid (( 1 R)-2-(2-naphthy I)- 1 -(( 1 R)-2-(2-naphthyl)- 1 -( 1 -phenethy I- 1 H- tetrazol-5-yl)ethyl-carbamoyl)ethyl)am ⁇ de, P ⁇ per ⁇ d ⁇ ne-4-carboxyl ⁇ c acid N-methyl-N-(( 1 R9-2-(2-naphthyl)- 1 -(( 1 R)2-(2-naphthyl)- 1 -thio- carbamoylethylcarbamoyl)ethyl)am ⁇ de,
  • H-A ⁇ b-H ⁇ s-D-2Nal-N-Me-D-Phe-Ser-NH 2 or the TFA salt thereof, (3-Am ⁇ nomethylbenzoyl)-D-2Nal-N-Me-D-Phe-Lys-NH 2 , or the TFA salt thereof, (4-P ⁇ per ⁇ d ⁇ necarbonyl)-D-2Nal-N-Me-D-Phe-NH 2 , or the TFA salt thereof,
  • growth hormone secretagogues are compounds disclosed in WO 94/13696, WO 94/19367, WO 95/14666, WO 94/11012, WO 96/15148, WO 95/34311 , WO 95/13069, WO 93/04081 and WO 97/07117, the contents of which are hereby incorporated by reference
  • the expression "increase the bone mass” is used to designate a condition wherein the balance between the resorption and formation of bone is shifted towards the formation so as to at least stop the loss of bone mass
  • a proposed dosage regimen for treating post-menopausal human beings having manifest osteoporosis which may e.g. be a spinal osteoporosis and a compression fracture of at least one vertebra or a manifest bone mineral density in lumbar spine of femoral neck, may e.g.
  • Kliogest® Novo Nordisk A/S, Bagsvaerd, Denmark; each tablet of Kliogest® comprises 2 mg of estradiol and 1 mg of norethisteron acetate) per day for two years and daily injections s.c.
  • GH neurotrophic factor
  • B-hGH Norditropin® biosynthetic human growth hormone
  • a somatostatin antagonist or GH secretagogue or a growth hormone component which can release the set amount of GH for a period of from about 2 days to about 28 days, preferably from about 2 days to about 14 days, more preferred from about 3 days to about 10 days, in particular from about 3 days to about 7 days, with intervals of from about 1 week to about 26 weeks, preferably from about 3 weeks to about 26 weeks, more preferably from about 6 weeks to about 12 weeks, more preferred from about 6 weeks to about 10 weeks, in particular about 8 weeks, between the periods in which of growth hormone is administered.
  • the injections may be carried out using a normal syringe or using a pen device such as a Nordiject® pen device (Novo Nordisk A/S, Bagsvaerd, Denmark).
  • the estrogen is preferably a preparation comprising a combination of an estrogen and a gestagen. For women who have no uterus, it is preferred to give an estrogen alone.
  • human growth hormone is used for the manufacture of a medicament for treating osteoporosis by simultaneous administration together with estrogen.
  • human growth hormone is used for the manufacture of a medicament for preventing the development of osteoporosis by simultaneous administration together with estrogen.
  • EXAMPLE 1 An approximately 62-year old postmenopausal woman with manifest osteoporosis was treated for one year with Kliogest® tablets orally every day and Norditropin® growth hormone subcutaneously for seven days every 8th week. Every day without interruption, one Kliogest® tablet was given In the seven-day periods when Norditropin® was administered, the dosage used was 0 2 lU/kg body weight/day. The osteoporosis was radiologically verified. The woman did not suffer from any other bone disease than osteoporosis. She had normal glucose metabolism and was not familiarly disposed to non-insulin dependent diabetes mellitus. She did not abuse alcohol or drugs and had no vaginal bleedings of unknown aetiology, and had never suffered thromboembolic disorders during estrogen treatment.
  • Bone mineral density of the lumbar spine (L2-L4), and of the femoral neck was assessed by quantitative digital radiography before the therapy started and after six and twelve months of therapy Bone mineral density at the spine increased by 3% and 15% after six and twelve months of therapy, respectively, compared to baseline.
  • Bone mineral density of lumbar spine had increased from 0.611 to 0.631 and 0.703, respectively after six and twelve months of combined estrogen and growth hormone therapy
  • Bone mineral density of the femoral neck increased by 11.4% after twelve month of therapy compared to baseline In g/cm 2 bone mineral density of the femoral neck had increased from 0.688 to 0 774
  • Kliogest® was administered daily in an oral form for 12 months.
  • GH was injected s.c. in the evening for 7 days every 2 months during the 12 months treatment period in a dose of 0.2 lU/mg/day.
  • the results in percentage change in BMD after 12 months treatment showed that the effect on BMD spine was greater in patients receiving the combined treatment of Kliogest® and GH, mean percent increase 9.5%, than in patients only treated with estrogen, mean percent increase 6.8%. No beneficial effect was seen in patients who received GH plus placebo or placebo alone.
  • the effect on BMD hip showed exactly the same dose-response relationship as in spine with a greater increase in patients receiving the combined treatment compared to either estrogen or GH alone.
  • Step A At 0 °C, ethyl chloroformate (1.10 mL, 11.5 mmol) was given dropwise to a solution of
  • Step B DMSO (1 22 mL, 17 2 mmol) was added to a solution of oxalyl chloride (1 1 mL, 12 9 mmol) at -78 °C in dichloromethane (15 L) The mixture was stirred for 15 mm at -78 °C A solution of 3-hydroxy-1 ,1-d ⁇ methylpropylcarbam ⁇ c acid tert-butyl ester (1 75 g, 8 6 mmol) in dichloromethane (10 mL) was added dropwise over a period of 15 mm The solution was stirred at -78 °C for another 15 mm Triethylamine (6 0 mL, 43 mmol) was added The solution was stirred at -78 °C for 5 m and then warmed to room temperature The solution was diluted with dichloromethane (100 L) and extracted with 1 N hydrochloric acid (100 mL) The aqueous phase was extracted with dichloromethane (50 mL) The combined
  • Step C Triethylphoshonoacetate (1.96 mL, 9.8 mmol) was dissolved in tetrahydrofuran (30 mL). Potassium tert-butoxide (1.10 g, 9.8 mmol) was added. The solution was stirred for 40 min at room temperature. A solution of 3-(tert-butoxycarbonylamino)-3-methylbutanal (1.10 g, 5.5 mmol) in Tetrahydrofuran (6 mL) was added. The solution was stirred at room temperature, for 75 min. It was diluted with ethyl acetate (100 mL) and 1N hydrochloric acid (100 mL). The phases were separated.
  • the aqueous phase was extracted with ethyl acetate (2 x 50 mL). T e combined organic phases were washed with saturated sodium hydrogen carbonate solution (60 mL) and dried over magnesium sulfate. The solvent was removed in vacuo.
  • the crude product was purified by column chromatography on silica (90 g) with ethyl acetate/hepatane (1 :4) to give 1.27 g of ethyl (2E)-5-(tert-butoxycarbonylamino)-5-methylhex-2-enoate.
  • Step D Ethyl (2E)-5-(tert-butoxycarbonylamino)-5-methylhex-2-enoate (1.233 g, 4.54 mmol) was dissolved in dioxane (20 mL). Lithium hydroxide (0.120 g, 5.00 mmol) was added as a solid. Water (10 mL) was added, until a clear solution was reached. The solution was stirred 16 h at room temperature. The solution was diluted with water (70 mL) and was extracted with tert-butyl methyl ether (2 x 100 mL).
  • Step E N-Tert-butoxycarbonyl-N-methyl-D-phenylalanine (1.22 g, 4.4 mmol), 1- hydroxybenzotriazole hydrate(0.59 g, 4.4 mmol) and 1-ethyl-3-(3-dimethyl- aminopropyl)carbodiimid hydrochloride (0.88 g, 4.6 mmol) were dissolved in N,N- dimethylformamide (25 mL) and stirred for 30 min. Methylamine (0.51 g of a 40% solution in methanol, 6.6 mmol) was added and the mixture was stirred overnight. Methylene chloride (80 mL) and water (100 mL) were added and the phases were separated.
  • Step F N-Methyl-N-((R)1-(methylcarbamoyl)-2-phenylethyl)carbamic acid tert-butyl ester (1.39 g, 7.23mmol) was dissolved in a mixture of trifluoroacetic acid (5 mL) and methylene chloride
  • Step G (R)-Tert-butoxycarbonyl-N-methylamino-3-(2-naphthyl)propionic acid (548 mg, 1.66 mmol) was dissolved in methylene chloride (5 mL); 1-hydroxy-7-azabenzotriazole (227 mg, 1,66 mmol) was added along with N,N-dimethylformamide (2 mL). 1-Ethyl-3-(3- dimethylaminopropyl)carbodiimide hydrochloride (351 mg, 1.83 mmol) was added and the solution was stirred for 15 min.
  • N-Methyl-2- methylamino-N-((1 R)-1 -(methylcarbamoyl)-2-phenylethyl)-3-(2-naphthyl)propionamide (332 mg, 0.82 mol) dissolved in methylene chloride (5 mL) and diisopropylethylamine (0.14 mL) were added and the mixture was stirred overnight under nitrogen atmosphere.
  • the mixture was diluted with methylene chloride (50 mL), washed with water (50 mL), sodium hydrogen carbonate (30 mL, saturated), and sodium hydrogensulfate (30 mL, 5%). The phases were separated and the organic phase was dried with magnesium sulfate and evaporated in vacuo.
  • the organic phase was extracted with a saturated aqueous solution of sodium hydrogen carbonate ( 50 mL). Citric acid (5 %) was added to the combined aqueous phases until pH 3, which were then extracted with ethyl acetate (2 x 50 mL) and the phases were separated.
  • the organic phase was washed with water (2 x 50 mL), an aqueous solution of sodium thiosulfate (5 %; 2 x 50 mL) and water (50 mL) and dried (magnesium sulfate). The solvent was removed in vacuo and the residue was dissolved in diethyl ether (10 mL). Dicyclohexylamine (10 mL) was added.

Abstract

A novel method of treating or preventing the outbreak of osteoporosis in an animal, e.g. a mammal, in particular a human being, comprises cyclic administration to the individual of a growth hormone component optionally supplemented with continuous administration of a medicament having estrogenic effect and/or a medicament having gestagenic effect.

Description

GROWTH HORMONE COMPONENT AND BONE ANΗ-RESORPTiVE AGENT IN CYCLIC (CO¬ HERENCE) TREATMENT OF OSTEOPOROSIS
FIELD OF THE INVENTION
The present invention relates to the treatment or prevention of osteoporosis
BACKGROUND OF THE INVENTION
Osteoporosis is a metabolic bone disease characterized by an absolute decrease in the bone mass Clinically this gives rise to an increased susceptibility to fractures Osteoporosis is a major problem in the developed countries
The growth of the human skeleton stops by the age of 20-30 years However, both cortical and trabecular bone is continuously renewed throughout life by the process of remodeling (see e g Parfitt, A M (1988) in Osteoporosis Etiology, diagnosis and management, B L Riggs, and L J Melton III, eds , Raven Press, New York) The remodeling sequence is initiated by osteoblasts or osteoblast-deπved cells (bone lining cells) digesting the endosteal membrane, thereby exposing the mineralized bone surface (Chambers, T J (1982), J Cell Sci , 52, 247-260) Osteoclasts are then recruited from marrow precursors and together with mononuclear cells they resorb a minute amount of bone digging out the resorption lacunae (Eriksen, E F et al , J Bone Min Res (1990), 5_, 311-319) which is later invaded by osteoblasts refilling the cavity with new bone In cortical bone, the end product of this process is the Haversian system (osteon) while in the trabecular bone it is seen as a trabecular osteon (Bone Structural Units. BSU or walls) The cells participating in the resorption and formation of one quantum of bone constitute the Bone Multicellular Unit (BMU) (Parfitt, A M et al , ibid) In normal subjects, a tight coupling between resorption and formation in both time and space tends to balance the two processes, thereby protecting against bone loss
The pathogenesis of osteoporosis is still largely unknown Recent histomorphometπc studies (Eriksen, E F , J Bone Mm Res (1990), 5, 311-319), however, have demonstrated a reduced osteoblastic activity in patients suffering from osteoporosis Eriksen et. al (Eriksen, E F ibid), investigated trabecular bone remodeling in 89 osteoporotic women (aged 66 ± 6 years), and found a highly significant reduction in mean wall thickness confirming an earlier report by Darby and Meunier (Calcif Tissue Int (1981), 33, 199-204) This reduction in mean thickness, leads to a pronounced imbalance between resorption and formation not found in subjects not suffering from osteoporosis
More than 40% of all post-menopausal women suffer low-energy fractures between the age of 50 and 70 years (Jensen, GF et al , Clin Orthop (1982), l≤S, 75-79) and the incidence of osteoporosis in males is increasing (Bengner, U et al , Calcif Tissue Int (1988), __Z, 293-296) Treatment with antiresorptive regimens is only able to provide for a 5 to 10% increase of the bone mass However, as the loss of bone in manifest osteoporosis often amounts from 30 to 50%, an optimal treatment should provide a corresponding increase in the bone mass
It is a commonly known method to treat menopausal women with estrogen in order to treat or prevent osteoporosis However, it is also known that administration of high doses of estrogen may increase the risk of developing cancer and does not provide a sufficiently large increase of the bone mass This has called for preparations comprising moderate amounts of estrogen increasing the demands on the efficiency in the use of the estrogen
It has been found that osteoblasts have receptors for both growth hormone and insulin growth factors I and II (IGF-I and IGF-II) (Bπxen et al , Potential Use of Growth Hormone in the Treatment of Osteoporosis, in Abstracts, Workshop on Growth No 5, September 25-26, 1992, Moltkes Palace, Copenhagen, Denmark) Although growth hormone has been found to increase bone mass considerably in rats and dogs, it does not seem to significantly affect bone mass in humans
The object of the present invention is to provide a more efficient method of treating or preventing the development of osteoporosis in humans which may ensure a more efficient use of estrogen in order to increase the bone formation over the level obtainable hitherto
SUMMARY OF THE INVENTION
The present invention relates, in a first aspect, to the treatment of osteoporosis in an animal and to the use of a growth hormone or a compound having growth hormone like or growth hormone releasing effect as well as somatostatin antagonists (in the following also collectively referred to in a generic way as the growth hormone component) and a composition with anti- resorptive action on bone (in the following referred to in a generic way as the estrogenic component) in the treatment
In another aspect, the invention relates to increasing the bone mass and preventing the development of osteoporosis in an animal and to the use of a growth hormone component and an estrogenic component in the treatment
Further details of the invention will appear from the appended claims
DETAILED DESCRIPTION OF THE INVENTION
It has surprisingly been found that when a growth hormone, somatostatin antagonist, or a growth hormone secretagogue (growth hormone component) is administered cyclically and, simultaneously, a composition with anti-resorptive action on bone is administered continuously to an animal, loss of the bone mass is stopped and an increase of bone mass is seen
The present invention relates to a method of treating as well as a method of preventing the development of osteoporosis and related disorders such as osteopenia in an animal compri¬ sing simultaneous administration to the animal of amounts of a growth hormone component and an estrogen component effective, in combination, to increase the bone mass in the animal
The term "animal" used in the present context includes, but is not limited to birds, such as chicken, ducks or turkeys, fish such as salmon, trout or tuna, and mammals such as cows, horses, sheep and human beings The animal to be treated is preferably a mammal and more preferred a human being, most preferably a woman, and the growth hormone is preferably human growth hormone
The term "simultaneous" used in connection with administration of the growth hormone component and the estrogen component in the present context is meant to designate administration of the two drugs in such a manner that, any time, both of them are available in adequate amounts to provide the benefit of the invention This does not imply that both drugs must be administered at exactly the same time as e g by precisely simultaneous injection of the two drugs, but rather that both drugs are intended to be given in any practical manner which provides for optimal availability thereof
As indicated above, it has turned out that, optimally, the growth hormone component is administered in a cyclic way which means that periods in which growth hormone component is administered alternate with periods where no growth hormone component is administered as further specified in the present text The estrogen component is preferably administered daily
One of the drugs may be administered by injection and the other may given orally, e.g in the form of a tablet Alternatively, it is envisaged that each medicament may, individually, be administered by injection, in the form of a patch for topical application, by nasal administration or in the form of a tablet Presently however, growth hormone is preferably administered by injection It could also be envisaged that growth hormone was used in a sustained release formulation
According to a preferred aspect of the invention, the estrogen is given continuously during the period of treatment whereas the growth hormone is given at intervals A preferred method of administration comprises daily administration of estrogen and penodical administration of growth hormone for a period of from about 2 days to about 28 days, preferably from about 2 days to about 14 days, more preferred from about 3 days to about 10 days, in particular from about 3 days to about 7 days, with intervals of from about 1 week to about 26 weeks, preferably from about 3 weeks to about 26 weeks, more preferably from about 6 to about 12 weeks, even more preferred from about 6 weeks to about 10 weeks, in particular about 8 weeks, between the periods in which of growth hormone is administered The estrogen component is administered in the amount from 0.001 mg to 10 mg/kg body weight/day When the estrogen component used is estradiol, the total amount given per day is from about 0.5 mg to about 4 mg, preferably from about 1 mg to about 2 mg. If another estrogen is used, an amount which has an equivalent anti-resorptive effect on bone is given When a gestagen is included in the regimen, this can be norethisterone acetate which is administered in an amount of from about 0 1 mg to about 2.0 mg, preferably from about 0.25 mg to about 2 0 mg per day according to the schedule If another estrogen is used, an equivalent amount is given
In the present context, "estrogen" is considered to encompass any preparation containing an estrogenic substance such as a natural human estrogen such as estrone, 17-β-estradιol and estradiol or derivatives thereof being cleaved in vivo to form natural estrogens, natural equine estrogens prepared from urine from horses, or artificial estrogens without steroid structure such as dienestrol The "estrogen" could also be selected from the group of non-steroidal estrogen directed therapeutics (NSERTs) such as Centchroman, Levormeloxifene, Raloxifene, Droloxifene, Tamoxifene, Idoxifene, or the like or structurally related compounds thereof, such as e g the compounds disclosed in WO 96/21656, WO 95/10513, US Patent 5,280,040, WO 96/09040, EP 0693488, WO 95/34557, EP 0617030, WO 93/10741, US Patent 5,254,568, EP 0683170, EP 0659413, EP 0652006, EP 0652007 and EP 0674903, the contents of which are hereby incorporated by reference As "estrogen" may be used a composition containing an estrogenic compound as the only pharmacologically active component or a composition which further to the estrogenic agent also contains a gestagen
In the present context, "growth hormone" may be growth hormone of any origin such as avian, bovine, equine, human, ovine, porcine, salmon, trout or tuna growth hormone, preferably bovine, human or porcine growth hormone, human growth hormone being most preferred The growth hormone used in accordance with the invention may be native growth hormone isolated from a natural source, e g by extracting pituitary glands in a conventional manner, or a growth hormone produced by recombinant techniques, e g as described in E.B Jensen and S Carlsen in Biotech and Bioeng 3S, 1-11 (1990) The growth hormone may also be a truncated form of growth hormone wherein one or more ammo acid residues has (have) been deleted, an analogue thereof wherein one or more ammo acid residues in the native molecule has (have) been substituted by another ammo acid residue, preferably a natural ammo acid residue, as long as the substitution does not have any adverse effect such as antigenicity or essentially reduced activity, or a derivative thereof, e g having an N- or C-terminal extension such as Met- hGH The preferred growth hormone is human growth hormone (hGH)
Compounds having growth hormone like or growth hormone releasing effect may e g be growth hormone releasing hormone (GHRH), growth hormone releasing factor or smaller oligo or polypeptides stimulating the release of growth hormone in vivo such as short-chain growth hormone releasing peptides, or growth factors such as IGF-I or IGF-ll, as well as somatostatin antagonists Examples of suitable growth hormone components are growth hormone (GH), IGF-I, IGF-ll, PACAP, GHRH, truncated GHRH, GHRP-1 , GHRP-2, GHRP-6, MK-677 disclosed in R P Margund et al , PNAS 92(15), 1995, p 7001, hexare n disclosed in WO 91/18016, as well as compounds disclosed in WO 95/17422, WO 95/17423, WO 96/05195, WO 96/22997, WO 96/24580, WO 96/24587, WO 97/00894, such compounds are e.g. H-Hιsψ(CH2NH)D-Trp-Ala-Trp-D-Phe-Lys-NH2, H-Hιs-D-Trpψ(CH2NH)Ala-Trp-D-Phe-Lys-NH2l H-Hιs-D-Trp-Alaψ(CH2NH)Trp-D-Phe-Lys-NH2, H-Hιs-D-Trp-Ala-Trpψ(CH2NH)D-Phe-Lys-NH2, H-His-D-Trp-Ala-Trp-D-Pheψ(CH2NH)Lys-NH2, H-D-Ala-D-2Nal-Alaψ(CH2NH)Trp-D-Phe-Lys-NH2, H-D-Ala-D-2Nal-Ala-Trp-D-Pheψ(CH2NH)Lys-NH2, (3-(4-lmιdazolyl)propιonyl)-D-2Nal-Ala-Trp-D-Pheψ(CH2NH)Lys-OH, (3-(4-lmιdazolyl)propιonyl)-D-2Nal-Ala-Trp-D-Pheψ(CH2NH)Lys-NH2, (3-(4-lmιdazolyl)acryloyl)-D-2Nal-Ala-Trp-D-Pheψ(CH2NH)Lys-NH2, H-D-Ala-D-Phe-Ala-Trp-D-Pheψ(CH2NH)Lys-NH2, (2R)-(H-D-Ala-D-Phe-Ala-Trp-NH)-3-phenylpropylamιne, (2S)-(H-D-Ala-D-2Nal-Alaψ(CH2NH)Trp-D-Phe-NH)-6-amιnohexanol, H-D-Ala-D-2Nal-Alaψ(CH2NH)Trp-D-Phe-NH2, 4-(H-D-Ala-D-2Nal-Alaψ(CH2NH)Trp-D-Phe-NH)butylamιne, (2R)-(H-D-Ala-D-2Nal-Ala-Trp-NH)-3-phenylpropylamιne, ((2R)-(H-D-Ala-D-2Nal-Ala-Trp-NH)-3-phenylproρylamιno)hexylamιne, (2R)-(H-D-2Nal-Ala-N-Bzl-Gly-NH)-3-phenylpropylamιne,
(2R)-(H-D-Ala-D-2Nal-Ala-N-Bzl-Gly-NH)-3-phenylpropylamιne, H-Aιb-D-2Nal-Ala-N-Bzl-Gly-D-Pheψ(CH2NH)Lys-NH2,
(2S)-((3-(4-lmιdazolyl)propιonyl)ψ(CH2NH)D-Phe-Ala-Tφ-D-Phe-NH)-6-amιnohexanol, (2S)-((3-(4-lmιdazolyl)propιonyl)-D-Pheψ(CH2NH)Ala-Trp-D-Phe-NH)-6-amιnohexanol, (2S)-((3-(4-lmιdazolyl)propιonyl)-D-Phe-Alaψ(CH2NH)Trp-D-Phe-NH)-6-amιnohexanol, (2S)-((3-(4-lmιdazolyl)propιonyl)-D-Phe-Ala-Trpψ(CH2NH)D-Phe-NH)-6-amιnohexanol, (2S)-(2R)-((3-(4-lmιdazolyl)propιonyl)-D-Phe-Ala-Trp-NH)-3-phenylpropylamιno)-6- ammohexanol,
3-((3-(4-lmιdazolyl)propιonyl)-D-Tφ-Ala-ψ(CH2NH)Trp-D-Phe-NH)propylamιne, (2S)-((3-(4-lmιdazolyl)propιonyl)-D-Phe-Ala-Trp-D-Pheψ(CH2NH)NH)-6-amιnohexanol, (2S)-((3-(4-lmιdazolyl)propιonyl)-D-Trp-Alaψ(CH2NH)Trp-D-Phe-NH)-6-amιnohexanol, 3-((3-(4-lmιdazolyl)propιonyl)-D-Trp-Alaψ(CH2NH)Trp-D-Phe-NH)propylamιne, H-D-Ala-D-2Nal-Ala-N-Bzl-Gly-D-Pheψ(CH2NH)Lys-NH2, H-Aιb-D-2Nal-Ala-N-Bzl-Gly-D-Pheψ(CH2NH2),
H-Ala-Hιsψ(CH2NH)D-2Nal-D-Phe-Lys-NH2,
H-Ala-Ala-D-2Nal-D-Phe-Lys-NH2,
H-Hιs-D-2Nal-D-Phe-Lys-NH2, (3-(4-lmιdazolyl)propιonyl)-D-2Nal-D-Phe-Lys-NH2,
H-D-Lys-D-2Nal-D-Phe-Lys-NH2,
H-5Apent-Hιs-D-2Nal-D-Phe-Lys-NH2,
H-D-Ala-D-2Nal-D-Phe-Lys-NH2,
H-5Apent-D-2Nal-D-Phe-Lys-NH2, (n-Propyl)-His-D-2Nal-D-Phe-Lys-NH2,
H-Ala-3Pyal-D-2Nal-D-Phe-Lys-NH2,
H-Ala-Phe(4-NH2)-D-2Nal-D-Phe-Lys-NH2,
H-D-Ala-His-D-2Nal-D-Phe-Lys-NH2,
(2-(4-lmιdazolyl)acetyl)-D-2Nal-D-Phe-Lys-NH2, (3-(4-lmidazolyl)acryloyl)-D-2Nal-D-Phe-Lys-NH2,
(3-Amιnomethyl benzoyl)-D-2Nal-D-Phe-Lys-NH2,
(3-Amιnophenylacetyl)-D-2Nal-D-Phe-Lys-NH2,
(4-Amιnophenylacetyl)-D-2Nal-D-Phe-Lys-NH2,
(3-Amιnocrotonoyl)-D-2Nal-D-Phe-Lys-NH2, (4-Pιperιdιno-carboxyl)-D-2Nal-D-Phe-Lys-NH2,
H-Ala-Hιs-D-2Nal-D-Phe-NH2,
(H-Ala-His-D-2Nal-D-Phe-NH)hexane,
6-(H-Ala-Hιs-D-2Nal-D-Phe-NH)hexylamιne,
5-(H-Ala-His-D-2Nal-D-Phe-NH)pentylanaιne, H-Ala-Hιs-D-2Nal-D-Pheψ(CH2NH)Lys-NH2,
H-Ala-Hιs-D-2Nal-D-Phe-Lys-OH,
(2S)-(H-Ala-His-D-2Nal-D-Phe-NH)-6-aminohexanol,
(2-(H-Ala-Hιs-D-2Nal-D-Phe-NH)ethyl)benzene,
2-(H-Ala-His-D-2Nal-D-Phe-NH)ethylamine, 4-((H-Ala-His-D-2Nal-D-Phe-NH)methyl)benzylamιne,
H-Ala-His-D-2Nal-D-Phe-Lys(maltosyl)-NH2,
H-Ala-His-D-2Nal-D-Phe-Phe-NH2,
H-Ala-His-D-2Nal-D-Phe-D-Phe-NH2,
H-Ala-Hιs-D-Phe-D-Phe-Lys-NH2l H-Ala-Hιs-D-Trp-D-Phe-Lys-NH2, H-Hιs-D-2Nal-D-Trp-Lys-NH2, H-Ala-Hιs-D-1 Nal-D-Phe-Lys-NH2, H-Ala-Phe-D-2Nal-D-Phe-Lys-NH2, H-Ala-Hιs-D-2Nal-D-Phe-Lys(maltosyl)-NH2,
(2R)-(H-Ala-Hιs-D-2Nal-D-Phe-Lys-NH)-3-phenylpropylamιne, H-Ala-N-Me-(2-amιnobenzoyl)-D-2Nal-D-Phe-Lys-NH2l (3-(Methylamιnomethyl)benzoyl)-D-2Nal-D-Phe-Lys-NH2, (4-(Ammomethyl)benzoyl)-D-2Nal-D-Phe-Lys-NH2, H-Hιs-Ala-D-2Nal-D-Phe-Lys-NH2,
4-(H-Ala-Hιs-D-2Nal-D-Phe-NH)butylamιne, 3-(H-Ala-Hιs-D-2Nal-D-Phe-NH)propylamιne, (3-(Dιmethylamιnomethyl)benzoyl)-D-2Nal-D-Phe-Lys-NH2, (3-Amιno-3-methylbutanoyl)-D-2Nal-D-Phe-Lys-NH2l (3-Amιnomethylbenzoyl)-D-hPhe-D-Phe-Lys-NH2l
(3-Amιnomethylbenzoyl)y(CH2NH)D-2Nal-D-Phe-Lys-NH2,
(3-Amιnomethylbenzoyl)-D-2Nal-D-hPhe-Lys-NH2,
(3-Amιno-3-methylbutanoyl)-Hιs-D-2Nal-D-Phe-Lys-NH2,
(3-Amιnomethylbenzoyl)-D-2Nal-N-Bzl-Gly-Lys-NH2, (2S)-(3-amιnomethylbenzoyl)y(CH2NH)-D-2Nal-D-Phe-NH)-6-amιnohexanol,
(2S)-((3-amιnomethylbenzoyl)-D-2Nal-D-Phe-NH)-6-amιnohexanol,
(3-Amιnomethylbenzoyl)-D-2Nal-D-Thιal-Lys-NH2,
(2S)-(H-Aιb-Hιsψ(CH2NH)-D-2Nal-D-Phe-NH)-6-amιnohexanol,
(3-Amιnomethylbenzoyl)-D-2Nal-D-3Pyal-Lys-NH2, (3-Amιnomethylbenzoyl)-D-2Nal-D-Phe(4-F)-Lys-NH2,
(3-Amιnomethylbenzoyl)-D-2Nal-D-Phe(4-OMe)-Lys-NH2,
(2-Amιnomethylphenylacetyl)-D-2Nal-D-Phe-Lys-NH2,
(2-Amιnomethylbenzoyl)-D-2Nal-D-Phe-Lys-NH2,
2-(H-Aιb-Hιs-D-2Nal-D-Phe-NH)-(4-pyndyl)ethane, H-Aιb-Phe-D-2Nal-D-Phe-Lys-NH2,
2-(H-Aιb-Hιs-D-2Nal-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane,
2-(H-Aιb-Hιs-D-2Nal-D-Phe-NH)-(4-pyπdyl)ethane,
H-Aιb-Hιsψ(CH2NH)-D-2Nal-D-Phe-Lys-OH,
(3-Amιnomethylbenzoyl)-D-2Nal-N-Me-D-Phe-Lys-NH2, H-Aιb-Hιs-D-2Nal-D-Phe-Gly-NH2,
H-Aιb-Hιs-D-2Nal-D-Phe-Ala-NH2,
H-Aιb-Hιs-D-2Nal-D-Phe-Orn-NH2
(5-Amιnomethylthιenyl-2-carbonyl)-D-2Nal-D-Phe-Lys-NH2, H-Aιb-Hιs-D-2Nal-D-Phe-D-Lys-NH2,
H-Aιb-Hιs-D-2Nal-D-Phe-Dab-NH2,
H-Aιb-Hιs-D-2Nal-D-Pheψ(CH2NH)-Lys-NH2l
H-Aιb-Hιs-N-Me-D-2Nal-D-Phe-Lys-NH2,
H-Aιb-Hιs-D-2Nal-D-Phe-N-Me-Lys-NH2, (3-Ammomethylthιenyl-2-carbonyl)-D-2Nal-D-Phe-Lys-NH2,
H-Aιb-Hιs-D-2Nal-N-Me-D-Phe-Lys-NH2,
H-Aιb-Hιs-D-2Nal-D-Phe-Lys-N(Me)2,
(3R)-Pιperιdιnecarbonyl-D-2Nal-D-Phe-Lys-NH2,
(3S)-Pιpendιnecarbonyl-D-2Nal-D-Phe-Lys-NH2l (3-Amιnomethylbenzoyl)-D-1 Nal-D-Phe-Lys-NH2,
H-Aιb-Hιs-D-2Nal-D-Trp-Lys-NH2,
(Furfuryl)-Aιb-Hιs-D-2Nal-D-Phe-Lys-NH2,
(2-Pyrιdylmethyl)-Aιb-Hιs-D-2Nal-D-Phe-Lys-NH2,
H-Aιb-(3-amιnomethylbenzoyl)-D-2Nal-D-Phe-Lys-NH2, H-Aιb-3Pyal-D-2Nal-D-Phe-Lys-NH2,
(3S)-Pιpeπdιnecarbonyl-D-2Nal-D-Phe-Lys-NH2l
(3R)-Pιpeπdιnecarbonyl-D-2Nal-D-Phe-Lys-NH2,
(2-(H-Aιb-Hιs-D-2Nal-NH)ethyl)benzene,
N,N-dι(2R-Hydroxypropyl)-(3-amιnomethylbenzoyl)-D-2Nal-D-Phe-Lys-NH2, (2R-Hydroxypropyl)-Aib-Hιs-D-2Nal-D-Phe-Lys-NH2,
(3-Amιnomethylbenzoyl)-D-2Nal-D-Pheψ(CH2NH)Lys-NH2,
(3-Amιnomethylbenzoyl)-N-Me-D-2Nal-D-Phe-Lys-NH2,
(3-Amιnomethylbenzoyl)-D-2Nal-D-Phe-N-Me-Lys-NH2,
H-D-Thr-Hιs-D-2Nal-D-Phe-Lys-NH2, H-Aιb-Hιs-D-2Nal-N-(phenethyl)-Gly-Lys-NH2,
(3-Amιnomethylbenzoyl)-D-2Nal-N-(phenethyl)-Gly-Lys-NH2, H-Hyp-Hιs-D-2Nal-D-Phe-Lys-NH2, H-Aιb-Hιs-N-Me-D-2Nal-N-(phenethyl)-Gly-Lys-NH2, H-Aιb-Hιs-N-Me-D-2Nal-N-Me-D-Phe-Lys-NH2, H-Aιb-Hιs-D-2Nal-D-Phey(CH2N(Me))Lys-NH2,
3-(H-Aιb-Hιs-D-2Nal-N-Me-D-Phe-NH)morpholιnopropane,
2-(H-Aιb-Hιs-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane,
(3R)-Pιpeπdιnecarbonyl-N-Me-D-2Nal-N-Me-D-Phe-Lys-NH2, 3-((Amιnomethylbenzoyl)-D-2Nal-N-Me-D-Phe-NH)morpholιnopropane,
2-(H-Aιb-Hιs-N-Me-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane,
2-(3R)-Pιperιdιnecarbonyl-N-Me-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane,
2-(3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)- ethane, 3-(H-Aιb-Hιs-N-Me-D-2Nal-N-Me-D-Phe-NH)morpholιnopropane,
3-((3R)-Pιperιdιnecarbonyl-N-Me-D-2Nal-N-Me-D-Phe-NH)morpholιnopropane,
3-((3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-NH)morpholιnopropane,
H-Aιb-Hιs-D-2Nal-N-Me-D-Phe-Hyp-NH2,
2-((3-Amιnomethylbenzoyl)-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane, 2-((3R)Pιperιdιnecarbonyl-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane,
H-Aιb-Hιs-D-2Nal-D-Phe-Lys-NH2,
3-Amιno-3-methyl-N-(4-oxo-5-(2'-(tetrazol-5-yl)bιphenyl-4-ylmethyl)-2,3,4,5-tetrahydro-1H- naphtho[2, 1 -b]azepιn-3-yl)butyramιde,
3-Amιno-3-methyl-N-(4-oxo-5-(4-(4-(5-methyl-[1 ,3,4]oxadιazol-2-yl)-thιen-3- yl)benzyl)-2,3,4,5-tetrahydro-IH-naphtho-[2,1-b]azepιn-3-yl)butyramιde,
3-((2R)-Hydroxypropylamιno)-3-methyl-N-(5-(4-(4-(5-methyl-[1 ,3,4]oxadιazol-2-yl)thιen-3- yl)benzyl)-4-oxo-2,3,4,5-tetrahydro-1 H-naphtho[2,1-b]azepιn-3-yl)butyramιde,
1-Amιnocyclopropanecarboxylιc acid (4-oxo-5-(2'-(1 H-tetrazole-5-yl)-bιphenyl-4-ylmethyl)-
2,3,4, 5-tetrahydro-1 H-naptho[2, 1 -b]azepιn-3-yl)amιde, 3-Amιno-3-methyl-N-(5-benzyl-4-oxo-2,3,4,5-tetrahydro-1H-naptho[2,1-b]azepιn-3- yl)butyramιde,
(3R) Pιpeπdιne-3-carboxylιc acid ((1R,2E)-4-hydroxymethyl-1-(2-naphthyl)methyl-5- phenylpent-2-enyl)amιde,
3-Amιnomethyl-N-((1R,2E)-4-hydroxymethyl-1-(2-naphthyl)methyl-5-phenylpent-2-enyl)- benzamide,
Pιperιdιne-4-carboxylιc acid (1-{[1-(3-carbamoyl-[1 ,2,4]oxadιazol-5-yl)-2-(2-naphthyl)ethyl]-N- methylcarbamoyl}-2-(2-naphthyl)ethyl) amide,
5-{(1R)-1-[(2R)-2-(Pιperιdιne-4-carbonylamιno)-3-(2-naphthyl)propιonyl-N-methylamιno]-2-(2- naphthyl)ethyl}-[1 ,2,4]oxadιazole-3-carboxylιc acid ethyl ester, 5- 1-[2-(3-Amιno ethylbenzoyl)-3-(2-naphthyl)propιonyl-N-methylamιno]-2-(2-naphthyl)ethyl}- [1 ,2,4]oxadιazol-3-carboxylιc acid ethyl ester,
5-{(1R)-1-[(2R)-2-(3-Amιnomethylbenzoylamιno)-3-(2-naphthyl)propιonylamιno]-2-phenyl- ethyl}-[1 ,2,4]oxadιazole-3-carboxylιc acid ethyl ester, or the tπflouroacetic acid salt thereof,
Pipeπdine 4-carboxylιc acid [(1 R)-1-{(1R)-1-(3-methyl-[1 ,2,4]oxadιazol-5-yl)-2-phenylethyl- carbamoyl}-2-(2-naphthyl)ethyi]amιde,
3-Amιnomethyl-N-[(1R)-1-{(1R)-1-(3-methyl-[1 ,2,4]oxadιazol-5-yl)-2-phenylethylcarbamoyl}-2- (2-naphthyl)ethyl]benzamιde, 4-Amιno-4-methyl-pent-2-enoιc acid [(1R)-1-{(1R)-1-(3-methyl-[1 ,2,4]oxadιazol-5-yl)-2-phenyi- ethylcarbamoyl}-2-(2-naphthyl)ethyl]amιde,
(3R)-Pιperιdιne 3-carboxyiιc acid [(1R)-1-((1R)-1-(3-methyl-[1 ,2,4]oxadιazol-5-yl)-2-phenylethyl- carbamoyl)-2-(2-naphthyl)ethyl]amιde,
3-Amιnomethyl-N-((1 R, 2E, 4S)-4-carbamoyl-5-(2-naphthyl)-1-(2-naphthyl)methylpent-2 -enyl)benzamιde
Pιperιdιne-4-carboxylιc acid ((1 R,2E,4S)-4-carbamoyi-5-(2-naphthyl)-1-(2-naphthyl)methylpent- 2-enyl) amide,
N-((1R)-1-(((1R)-1-(((1S)-5-Amιno-1-(dιmethylcarbamoyl)pentylcarbamoyl)-2-phenylethoxy)- methyl)-2-(2-naphthyl)ethyl)-3-amιnomethylbenzamιde, N-((1 R,4S)-4-(((1S)-5-Amιno-1-(dιmethylcarbamoyl)pentyl)carbamoyl)-1-((2-naphthyl)methyl)- 2-oxo-5-phenylpentyl)-3-amιnomethylbenzamιde,
N-((1R,2R,4S)-4-(((1S)-5-Amιno-1-(dιmethylcarbamoyl)pentyl)carbamoyl)-2-hydroxy-1- (2-naphthyl)methyl-5-phenylpentyl)-3-amιnomethylbenzamιde,
Pιpendιne-3-carboxylιc acid ((1R, 2R, 4S)-4-(((1S)-5-amιno-1-(dιmethylcarbamoyl)pentyl)- carbamoyl)-2-hydroxy-1-((2-naphthyl)methyl)-5-phenylpentyl) amide,
5-((1R)-1-(N-Methyl-N-((2R)-3-(2-naphthyl)-2-(pιperιdιn-4-yl-carbonylamιno)propιonyl)amιno)-2- (2-naphthyl)ethyl)-[1 ,2,4]oxadιazole-3-carboxylιc acid ethyl ester,
5-((1R)-1-(N-((2R)-2-(3-Amιnomethylbenzoylamιno)-3-(2-naphthyl)propιonyl)-N-methylamιno)- 2-(2-naphthyl)ethyl)-[1,2,4]oxadιazole-3-carboxylιc acid ethyl ester, 5-((1R)-1-(N-((2R)-2-(3-Amιnomethylbenzoylamιno)-3-(2-naphthyl)propιonyl)-N-methylamιno)- 2-phenylethyl)-[1 ,3,4]oxadιazole-2-carboxylιc acid amide,
(2E)-5-Amιno-5-methylhex-2-enoιc acid {(1 R)-1 -[N-methyl-N-(( 1 R)-1 -(3-methyl-[1 ,2,4]- oxadιazol-5-yl)-2-(2-naphthyl)-ethyl)carbamoyl]-2-(2-naphthyl)ethyl} amide, 4-Amιno-4-methylpent-2-enoιc acid N-[(1R)-1-{N-methyl-N-[(1R)-1-(3-methyl-[1 ,2,4]oxadιazol- 5-yl)-2-(2-naphthyl)ethyl]carbamoyl}-2-(2-naphthyl)ethyl]-N-methylamιde, 4-Amιno-4-methylpent-2-enoιc acid [(1 R)-1-{N-methyl-N-[(1 R)-1-(3-methyl-[1 ,2,4]oxadιazol-5- yl)-2-(2-naphthyl)ethyl]carbamoyl}-2-(2-naphthyl)ethyl]amιde, 3-Amιnomethyl-N-(( 1 R)-1 -{N-[( 1 R)-1 -(((dιmethylcarbamoyl)methoxy)methyl)-2-phenylethyl]-N- methylcarbamoyl}-2-(2-naphthyl)ethyl)-N-methylbenzamιde,
5-((1R)-1-(((2R)-2-(((4E)-4-Amιno-4-methylpent-2-enoyl)methylamιno)-3-(2-naphthyl)- propιonyl)methylamιno)-2-phenylethyl)-[1 ,3,4]-oxadιazole-2-carboxylιc acid amide, Pιperιdιne-4-carboxylιc acid N-methyl-N-{-1([methyl-1-(3-methyl-[1 ,2,4]oxadιazole-5-yl)-2-(2- naphthyl)ethylcarbamoyl)-2-(2-naphthyl)ethyl}amιde,
Pιpeπdιne-4-carboxylιc acid N-{-1([methyl-1-(3-methyl-[1 ,2,4]-oxadιazole-5-yl)-2-(2- naphthyl)ethylcarbamoyl)-2-(2-naphthyl)ethyl}amιde,
5-{1-[2-(pιperιdιne-4-carbonylamιno)-3-(2-naphthyl)propιonyl-N-methylamιno]-2-(2- naphthyl)ethyl}-[1 ,2,4]oxadιazole-3-carboxylιc acid 2-propyl ester,
5-{1-[2-(pιpeπdιne-4-carbonylamιno)-3-(2-naphthyl)propιonyl-N-methylamιno]-2-(2- naphthyl)ethyl}-[1 ,2,4]oxadιazole-3-carboxylιc acid, tπfluoro acetate,
Pιperιdιne-4-carboxylιc acid (1-{[1-(3-methylcarbamoyl-[1 ,2,4]oxadιazol-5-yl)-2-(2-naphthyl)- ethyl]-N-methylcarbamoyl}-2-(2-naphthyl)ethyl)amιde, (2E)-5-Amιno-5-methylhex-2-enoιc acid {1-[N-(1-(3-benzylcarbamoyl-[1 ,2,4]oxadιazol-5-yl)-2- phenylethyl)-N-methyl-carbamoyl]-2-(2-naphthyl)ethyl} amide,
(2E)-5-Amιno-5-methylhex-2-enotc acid N-{(1R)-1-[((1R)-1-benzyl-2,5-dιhydroxypentyl)-N- methylcarbamoyl]-2-(2-naphthyl)ethyl}-N-methylamιde,
3-Amιnomethyl-N-((1 R)-1 -{N-[(1 R)-1 -(2-hydroxyethoxymethyl)-2-phenylethyl]-N-methyl- carbamoyl}-2-(2-naphthyl)ethyl)-N-methylbenzamιde,
Pιperιdιne-4-carboxylιc acid ((1 R,2E)4-hydroxymethyl-5-(2-naph-thyl)-1 -((2-naphthyl)methyl)- pent-2-enyl)amιde,
Pιpeπdιne-4-carboxylιc acid (( 1 R)-2-(2-naphthy I)- 1 -(( 1 R)-2-(2-naphthyl)- 1 -( 1 -phenethy I- 1 H- tetrazol-5-yl)ethyl-carbamoyl)ethyl)amιde, Pιperιdιne-4-carboxylιc acid N-methyl-N-(( 1 R9-2-(2-naphthyl)- 1 -(( 1 R)2-(2-naphthyl)- 1 -thio- carbamoylethylcarbamoyl)ethyl)amιde,
Pιperιdιne-4-carboxylιc acid ((1 R)-1 -((1 R)-1-(4-carbamoyl-5-phe-nyl-1 ,3-thιazol-2-yl)-2-(2- naphthyl)ethylcarbamoyl)-2-(2-naphthyl)ethyl)amιde, (2E)-5-Amιno-5-methylhex-2-enoιc acid {1-[N-(1-(3-methylcarbamoyl-[1 ,2,4]oxadιazol-5-yl)-2- phenylethyl)-N-methyl-carbamoyl]-2-(2-naphthyl)ethyl} amide,
(2E)-5-Amιno-5-methylhex-2-enoιc acid {1-[N-(1-(3-dιmethylcarbamoyl-[1 ,2,4]oxadιazol-5-yl)-2- phenylethyl)-N-methyl-carbamoyl]-2-(2-naphthyl)ethyl} amide, (2E)-5-Amιno-5-methyl-N-((1 R)-1-(N-((1 R)-1-(2-hydroxyethoxymethyl)-2-phenylethyl)-N- methylcarbamoyl)-2-(2-naphthyl)ethyl)-N-methylhex-2-enoιc acιd amide,
(2E)-5-Amιno-5-methyl-N-((1 R)-1-(N-((1R)-1-(2-hydroxy-2-methylpropoxymethyl)-2- phenylethyl)-N-methylcarbamoyl)-2-(2-naphthyl)ethyl)-N-methylhex-2-enoιc acιd,
1-Benzyl-3-(3-(morpholιn-4-yl)propyl)-1-(naphth-2-yl)methyt-thιourea, or the hydrchlonde salt thereof
1-Benzyl-3-(3-dιmethylamιnopropyl)-1-phenyl-thιourea, or the hydrochloride salt thereof,
2-[3-(3-(Morpholιn-4-yl)propyl)-1-(naphth-2-yl)methyl-thιoureιdo]-3-phenyl-propιonamιde,
N-(4-Amιnobutyl)-2-[3-((3-amιno-3-methyl)butyl)-1-(naphth-2-yl)methyl-thιoureιdo]-3-phenyl- propionamide, N-(4-Amιnobutyl)-2-(N-(naphth-2-yl)methyl-N'-(pιpeπdιn-3-yl)methyl-guanιdιno)-3-phenyl- propioπamide,
N-(4-Amιnobutyl)-2-[1-methyl-3-(naphth-2-yl)methyl-3-(2-(pιpeπdιn-2-yl)ethyl)-thιoureιdo]-3-
(naphth-2-yl)propιonamιde,
3-(3-(Morpholιn-4-yl)propyl)-1-(naphth-2-yl)methyl-1-[2'-(1 H-tetrazol-5-yl)-bιphenyl-4- ylmethyl]-thιourea, or the hydrochloride salt thereof,
N-((1-Carbamoyl-2-phenyl)ethyl-N-methyl-2-[3-((3-morpholιn-4-yl)propyl)- thιoureιdo]-3-
(naphth-2-yl)propιonamιde,
3-(3-(Dιmethylamιno)propyl)-1-(naphth-1-yl)methyl-1-phenylthιourea, or the hydrochloride salt thereof, 1-Benzyl-3-(3-(moφholιn-4-yl)propyl)-1-phenylthιourea,
1 ,1-Dιbenzyl-3-(3-(morpholιn-4-yl)propyl)thιourea, or the hydrochloride salt thereof,
1-Benzyl-3-(3-(dιmethylamιno)propyl)-1-((naphth-2-yl)methyl)thιourea, or the hydrochloride salt thereof,
1-Benzyl-3-(3-(morpholιn-4-yl)propyl)-1-(pheneth-2-yl)thιourea, or the hydrochloride salt thereof,
1-Benzyl-3-(3-(moφholιn-4-yl)propyl)-1-(quιnolιn-3-yl)thιourea, or the dihydrochloπde salt thereof,
1-Benzyl-3-(3-(morpholιn-4-yl)propyl)-1-(pyndιn-2-yl)thιourea, or the hydrochloride salt thereof, 1-Benzyl-3-(3-(morpholιn-4-yl)propyl)-1-(4-methoxyphenyl)thιourea, or the hydrochloride salt thereof,
1-Benzyl-3-(3-(morpholιn-4-yl)propyl)-1-(4-([1 ,2,3]thιadιazol-4-yl)benzyl)thιourea, or the hydrochloride salt thereof, or 3-(3-Dιmethylamιnopropyl)-1-((naphth-2-yl)methyl)-1-phenylthιourea, or the hydrochloride salt thereof,
2-Phenyl-3,4-dιhydro-2H-quιnolιne-1-carbothιoιc acid (3-(dιmethylamιno) propyl) amide, or the hydrochloride salt thereof,
Benzyl-3,4-dιhydro-2H-quιnolιne-1-carbothιoιc acid (3-(dιmethylamιno)propyl)amιde, 1-(3-((Morpholιn-4-yl)propyl)thιocarbamoyl)-1 ,2,3,4-tetrahydro-1H-quιnolιne-2-carboxylιc acιd
N-(1 -carbamoyl-2-(napht-1 -yl)ethyl)-N-methylamιde,
2-(3-((Morpholιn-4-yl)propyl)thιocarbamoyl)-1 ,2,3,4-tetrahydro-1H-ιsoquιnolιne-3-carboxylιc acid N-[1-((4-amιnobutyl)carbamoyl)-2-(naphth-1-yl)ethyl]-N-methylamιde, or
2-Phenyl-3,4-dιhydro-2H-quιnoiιne-1-carbothιoιc acιd (3-(morpholιn-4-yl)propyl) amide, or the hydrochloride salt thereof,
(R)-2-((3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me)-3-phenylpropanol, or the TFA salt thereof,
3-((3-Amιnomethylbenzoyl))N-Me-D-2Nal-N-Me-D-Phe-NH)-1-1N,N-dιmethylamιnopropane, or the TFA salt thereof,
3-(((3R)-3-Pιperιdιnecarbonyl)-N-Me-D-2Nal-N-Me-D-Phe-NH)-1-N,N-dιmethylamιnopropane, or the TFA salt thereof,
2-(((3R)-3-Pιperιdιnecarbonyl)-N-Me-D-2Nal-N-Me-D-Phe-NH)-(1-methyl-2-pyrrolιdιnyl)ethane, or the TFA salt thereof,
H-Aιb-Hιs-D-2Nal-N-Me-D-Phe-Ser-NH2, or the TFA salt thereof, (3-Amιnomethylbenzoyl)-D-2Nal-N-Me-D-Phe-Lys-NH2, or the TFA salt thereof, (4-Pιperιdιnecarbonyl)-D-2Nal-N-Me-D-Phe-NH2, or the TFA salt thereof,
((3R)-3-Pιperιdιnecarbonyl)-D-2Nal-N-Me-D-Phe-NH2, or the TFA salt thereof, (3-Amιnomethylbenzoyl)-D-Phe-N-Me-D-Phe-NH2, or the TFA salt thereof, (3-Ammomethylbenzoyl)-N-Me-D-Phe-N-Me-D-Phe-Lys-NH2, or the TFA salt thereof, ((3R)-3-Pιperιdιnecarbonyl)-N-Me-D-Phe-N-Me-D-Phe-Lys-NH2, or the TFA salt thereof, H-Aιb-Hιs-N-Me-D-Phe-N-Me-D-Phe-Lys-NH2, or the TFA salt thereof,
((3R)-3-Pιpeπdιnecarbonyl)-N-Me-D-2Nal-N-Me-D-Phe-NH2, or the TFA salt thereof,
(2R)-2-((3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me)-3-(2-naphthyl)propanol, or the TFA salt thereof,
(3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-NH2 , or the TFA salt thereof, 3-((3-Amιnomethylbenzoyl)-N-Me-D-Phe-NH)-1-N,N-dιmethylamιnopropane,
H-Aιb-Hιs-N-Me-D-2Nal-N-Me-D-Phe-NH2, or the TFA salt thereof,
(3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-Lys-NH2,
H-Aιb-Ala-D-2Nal-N-Me-D-Phe-Lys-NH2, or the TFA salt thereof, H-Aιb-Hιs-D-2Nal-N-Me-D-Phe-NH2, or the TFA salt thereof,
2-((3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-NH)-1-morpholιnoethane,
(3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-NH-Me,
3-((3-Methylammomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-NH)-1-N,N-dιmethyl- aminopropane, (3-Amιnomethylbenzoyl)-N-Me-D-2Nal-N-Me-D-Phe-N-Me2,
H-Aιb-Hιs-N-Me-D-2Nal-N-Me-D-Phe-NH2,
3-Amιnomethylbenzoyl-N-Me-D-2Nal-N-Me-D-Phe-NH-CH3, or the TFA salt thereof,
3-methylamιnomethylbenzoyl-N-Me-D-2Nal-N-Me-D-Phe-NH-CH3, or the TFA salt thereof,
H-Aιb-Hιs-N-Me-D-2Nal-N-Me-D-Phe-NHMe, or the HCI salt thereof, and Pιperιdιne-4-carboxylιc acιd-N-((1R)-1-(N-((1R)-2-(4-ιodophenyl)-1-
(methylcarbamoyl)ethyl)-N-methylcarbamoyl)-2-(2-naphthyl)ethyl)-N-methylamιde,
(2E) 5-Amιno-5-methylhex-2-enoιc acid N-methyl-N-((1R)-1-(N-methyl-N-((1R)-1-(methyl- carbamoyl)-2-phenylethyl)carbamoyl)-2-(2-naphthyl)ethyl)amιde, or the hydrochloride salt thereof, (2E)-5-Amιno-5-methylhex-2-enoιc acid N-((1 R)-1-(((1 R)-1-((2S)-2-hydroxypropylcarbamoyl)-2- phenylethyl)-methylcarbamoyl)-2-(2-naphthyl)ethyl)-N-methylamιde, or
(2E)-5-Amιno-5-methylhex-2-enoιc acid ((1R)-1-(((1R)-2-(4-fluorophenyl)-1-(methylcarbamoyl)- ethyl)methylcarbamoyl)-2-(2-naphthyl)ethyl)methylamιde
Other examples of suitable growth hormone secretagogues are compounds disclosed in WO 94/13696, WO 94/19367, WO 95/14666, WO 94/11012, WO 96/15148, WO 95/34311 , WO 95/13069, WO 93/04081 and WO 97/07117, the contents of which are hereby incorporated by reference
For the purpose of the present invention, the expression "increase the bone mass" is used to designate a condition wherein the balance between the resorption and formation of bone is shifted towards the formation so as to at least stop the loss of bone mass A proposed dosage regimen for treating post-menopausal human beings having manifest osteoporosis, which may e.g. be a spinal osteoporosis and a compression fracture of at least one vertebra or a manifest bone mineral density in lumbar spine of femoral neck, may e.g. be treatment with one tablet of Kliogest® (Novo Nordisk A/S, Bagsvaerd, Denmark; each tablet of Kliogest® comprises 2 mg of estradiol and 1 mg of norethisteron acetate) per day for two years and daily injections s.c. of 0.01-1 IU per kg body weight per day, preferably 0.1-0.2 IU per kg body weight per day, more preferably 0.2 IU per kg body weight per day (the specific activity of GH is 3 lU/mg), of Norditropin® biosynthetic human growth hormone (B-hGH, Novo Nordisk A/S, Bagsvaerd, Denmark), a somatostatin antagonist or GH secretagogue or a growth hormone component which can release the set amount of GH for a period of from about 2 days to about 28 days, preferably from about 2 days to about 14 days, more preferred from about 3 days to about 10 days, in particular from about 3 days to about 7 days, with intervals of from about 1 week to about 26 weeks, preferably from about 3 weeks to about 26 weeks, more preferably from about 6 weeks to about 12 weeks, more preferred from about 6 weeks to about 10 weeks, in particular about 8 weeks, between the periods in which of growth hormone is administered. The injections may be carried out using a normal syringe or using a pen device such as a Nordiject® pen device (Novo Nordisk A/S, Bagsvaerd, Denmark). The estrogen is preferably a preparation comprising a combination of an estrogen and a gestagen. For women who have no uterus, it is preferred to give an estrogen alone.
According to another aspect of the invention, human growth hormone is used for the manufacture of a medicament for treating osteoporosis by simultaneous administration together with estrogen.
According to yet another aspect of the invention, human growth hormone is used for the manufacture of a medicament for preventing the development of osteoporosis by simultaneous administration together with estrogen.
The invention is further illustrated in the following example which is not in any way intended to limit the scope of the invention as claimed.
EXAMPLE 1: An approximately 62-year old postmenopausal woman with manifest osteoporosis was treated for one year with Kliogest® tablets orally every day and Norditropin® growth hormone subcutaneously for seven days every 8th week. Every day without interruption, one Kliogest® tablet was given In the seven-day periods when Norditropin® was administered, the dosage used was 0 2 lU/kg body weight/day. The osteoporosis was radiologically verified. The woman did not suffer from any other bone disease than osteoporosis. She had normal glucose metabolism and was not familiarly disposed to non-insulin dependent diabetes mellitus. She did not abuse alcohol or drugs and had no vaginal bleedings of unknown aetiology, and had never suffered thromboembolic disorders during estrogen treatment.
Bone mineral density of the lumbar spine (L2-L4), and of the femoral neck was assessed by quantitative digital radiography before the therapy started and after six and twelve months of therapy Bone mineral density at the spine increased by 3% and 15% after six and twelve months of therapy, respectively, compared to baseline. In g/cm2 bone mineral density of lumbar spine had increased from 0.611 to 0.631 and 0.703, respectively after six and twelve months of combined estrogen and growth hormone therapy
Bone mineral density of the femoral neck increased by 11.4% after twelve month of therapy compared to baseline In g/cm2 bone mineral density of the femoral neck had increased from 0.688 to 0 774
EXAMPLE 2.
A randomized double-blind placebo controlled study with four arms in 57 women (aged 45-75 years) with post-menopausal osteoporosis (BMD > 2SD below mean, and one spinal fracture(s) and/or colles fracture(s)), has been performed
Four arms design:
GH + Kliogest®
GH + Placebo Placebo + Kliogest®
Placebo + Placebo.
Kliogest® was administered daily in an oral form for 12 months. GH was injected s.c. in the evening for 7 days every 2 months during the 12 months treatment period in a dose of 0.2 lU/mg/day. The results in percentage change in BMD after 12 months treatment showed that the effect on BMD spine was greater in patients receiving the combined treatment of Kliogest® and GH, mean percent increase 9.5%, than in patients only treated with estrogen, mean percent increase 6.8%. No beneficial effect was seen in patients who received GH plus placebo or placebo alone.
The effect on BMD hip showed exactly the same dose-response relationship as in spine with a greater increase in patients receiving the combined treatment compared to either estrogen or GH alone.
The following three examples illustrate preferred GH secretagogues.
EXAMPLE 3:
(2E) 5-Amino-5-methylhex-2-enoic acid N-methyl-N-((1R)-1-(N-methyl-N-((1R)-1-(methyl- carbamoyl)-2-phenylethyl)carbamoyl)-2-(2-naphthyl)ethyl)amide hydrochloride:
Figure imgf000020_0001
3-Hydroxy-1 , 1 -dimethylpropylcarbamic acid tert-butyl ester:
Figure imgf000020_0002
Step A: At 0 °C, ethyl chloroformate (1.10 mL, 11.5 mmol) was given dropwise to a solution of
3-tert-butoxycarbonylamino-3-methylbutanoic acid (2.50 g, 11.5 mmol) and triethylamine (1.92 mL, 13.8 mmol) in tetrahydrofuran (10 mL). The solution was stirred for 40 min at 0 βC. The formed precipitate was filtered off and washed with tetrahydrofuran (20 mL) The liquid was immediately cooled to 0 °C A 2M solution of lithium boronhydπde in tetrahydrofuran (144 mL, 28 8 mmol) was added dropwise The solution was stirred at 0 °C for 2 h, and then warmed to room temperature over a period of 4 h It was cooled to 0 °C Methanol (5 mL) was added carefully 1N Hydrochloric acid (100 mL) was added The solution was extracted with ethyl acetate (2 x 100 mL, 3 x 50 mL) The combined organic layers were washed with saturated sodium hydrogen carbonate solution (100 mL) and dried over magnesium sulfate The solvent was removed in vacuo The crude product was chromatographed on silica (110 g) with ethyl acetate/heptane 1 2 to give 1 84 g of 3-hydroxy-1 ,1-dιmethylpropylcarbamιc acid tert-butyl ester
Η-NMR (CDCy d 1 33 (s, 6 H), 1 44 (s, 9 H), 1 88 (t, 2 H), 1 94 (br, 1 H), 3 75 (q, 2 H), 4 98 (br, 1 H)
3-(tert-Butoxycarbonylamιno)-3-methylbutanal
Figure imgf000021_0001
Step B. DMSO (1 22 mL, 17 2 mmol) was added to a solution of oxalyl chloride (1 1 mL, 12 9 mmol) at -78 °C in dichloromethane (15 L) The mixture was stirred for 15 mm at -78 °C A solution of 3-hydroxy-1 ,1-dιmethylpropylcarbamιc acid tert-butyl ester (1 75 g, 8 6 mmol) in dichloromethane (10 mL) was added dropwise over a period of 15 mm The solution was stirred at -78 °C for another 15 mm Triethylamine (6 0 mL, 43 mmol) was added The solution was stirred at -78 °C for 5 m and then warmed to room temperature The solution was diluted with dichloromethane (100 L) and extracted with 1 N hydrochloric acid (100 mL) The aqueous phase was extracted with dichloromethane (50 mL) The combined organic layers were washed with saturated sodium hydrogen carbonate solution (100 mL) and dried over magnesium sulfate The solvent was removed in vacuo The crude product was punfied by column chromatography on silica (140 g) with ethyl acetate/heptane (1 3) to give 1 10 g of 3- (tert-butoxycarbonylamιno)-3-methyibutanal
MHz-Η-NMR (CDCI3) d 1 39 (s, 6 H), 1 45 (s, 9 H), 2 85 (d, 2 H), 4 73 (br 1 H), 9 80 (t, 1 H) Ethyl (2E)-5-(tert-Butoxycarbonylamino)-5-methylhex-2-enoate:
Figure imgf000022_0001
Step C: Triethylphoshonoacetate (1.96 mL, 9.8 mmol) was dissolved in tetrahydrofuran (30 mL). Potassium tert-butoxide (1.10 g, 9.8 mmol) was added. The solution was stirred for 40 min at room temperature. A solution of 3-(tert-butoxycarbonylamino)-3-methylbutanal (1.10 g, 5.5 mmol) in Tetrahydrofuran (6 mL) was added. The solution was stirred at room temperature, for 75 min. It was diluted with ethyl acetate (100 mL) and 1N hydrochloric acid (100 mL). The phases were separated. The aqueous phase was extracted with ethyl acetate (2 x 50 mL). T e combined organic phases were washed with saturated sodium hydrogen carbonate solution (60 mL) and dried over magnesium sulfate. The solvent was removed in vacuo. The crude product was purified by column chromatography on silica (90 g) with ethyl acetate/hepatane (1 :4) to give 1.27 g of ethyl (2E)-5-(tert-butoxycarbonylamino)-5-methylhex-2-enoate.
'H-NMR (CDCI3): d 1.30 (s, 6 H); 1.30 (t, 3 H); 1.46 (s, 9 H); 2.62 (d, 2 H); 4.27 (q, 2 H); 4.42 (br, 1 H); 5.88 (d, 1 H); 6.94 (td, 1 H).
(2E)-5-(tert-Butoxycarbonylamino)-5-methylhex-2-enoic acid:
Figure imgf000022_0002
Step D: Ethyl (2E)-5-(tert-butoxycarbonylamino)-5-methylhex-2-enoate (1.233 g, 4.54 mmol) was dissolved in dioxane (20 mL). Lithium hydroxide (0.120 g, 5.00 mmol) was added as a solid. Water (10 mL) was added, until a clear solution was reached. The solution was stirred 16 h at room temperature. The solution was diluted with water (70 mL) and was extracted with tert-butyl methyl ether (2 x 100 mL). The aqueous phase was acidified with 1N sodium hydrogensulfate solution (pH = 1) and was extracted with tert-butylmethylether (3 x 70 mL). The organic phases were combined and dried over magnesium sulfate. The solvent was removed in vacuo to give 1.05 g of (2E)-5-(tert-butoxycarbonylamino)-5-methylhex-2-enoic acid. The crude product was used for further syntheses. Η-NMR (DMSO dβ): d 1.15 (s, 6 H); 1.35 (s, 9 H); 2.53 (d, 2 H); 5.75 (d, 1 H); 6.57 (br, 1 H); 6.75 (td, 1 H); 12.15 (s, 1 H).
N-Methyl-N-((R)-1-(methylcarbamoyl)-2-phenylethyl)carbamic acid tert-butyl ester:
Figure imgf000023_0001
Step E: N-Tert-butoxycarbonyl-N-methyl-D-phenylalanine (1.22 g, 4.4 mmol), 1- hydroxybenzotriazole hydrate(0.59 g, 4.4 mmol) and 1-ethyl-3-(3-dimethyl- aminopropyl)carbodiimid hydrochloride (0.88 g, 4.6 mmol) were dissolved in N,N- dimethylformamide (25 mL) and stirred for 30 min. Methylamine (0.51 g of a 40% solution in methanol, 6.6 mmol) was added and the mixture was stirred overnight. Methylene chloride (80 mL) and water (100 mL) were added and the phases were separated. The organic phase was washed with sodium hydroxide (20 mL, 1N), sodium hydrogβnsulfate (50 mL, 10 %) and water (50 mL). The organic phase was dried (magnesium sulfate) and the solvent removed in vacuo to afford 1.39 g of N-methyl-N-((R)1-(methylcarbamoyl)-2-phenylethyl)carbamic acid tert-butyl ester.
Η-NMR (CDCI3): d 1.25, 1.35 (two s (br), 9H); 2.73-2.94 (m, 7H); 3.30-3.50 (m, 1H); 4.68, 4.90 (two m, 1H); 5.90, 6.12 (two s (br); 1H); 7.12-7.25 (m, 5H).
(R)-N-Methyl-2-methylamino-3-phenylpropionamide:
Figure imgf000023_0002
Step F: N-Methyl-N-((R)1-(methylcarbamoyl)-2-phenylethyl)carbamic acid tert-butyl ester (1.39 g, 7.23mmol) was dissolved in a mixture of trifluoroacetic acid (5 mL) and methylene chloride
(10 mL) and stirred for 45 min. The volatiles were removed in vacuo and the residue was stirred with a mixture of ethyl acetate (100 mL) and water (100 L). Sodium hydrogen carbonate (50 mL, saturated) was added and the phases were separated. The organic phase was dried (magnesium sulfate) and the solvent removed in vacuo to afford 330 mg of (R)-N- methyl-2-methylamino-3-phenylpropionamide.
Η-NMR (CDCI3): d 2.1 (s(br), 3H); 2.32 (s, 3H); 2.77 (dd, 1H); 2.81 (two s, 3H); 3.21 (dd, 1H); 3.32 (dd, 1H); 7.12 (s(br), 1H); 7.20-7.34 (m, 5H).
N-Methyl-N-{(1 R)-1-(N-methyl-N-((1 R)-1 -(methylcarbamoyl)-2-phenylethyl)carbamoyl)-2-(2- naphthyl)ethyl}carbamic acid tert-butyl ester:
Figure imgf000024_0001
Step G: (R)-Tert-butoxycarbonyl-N-methylamino-3-(2-naphthyl)propionic acid (548 mg, 1.66 mmol) was dissolved in methylene chloride (5 mL); 1-hydroxy-7-azabenzotriazole (227 mg, 1,66 mmol) was added along with N,N-dimethylformamide (2 mL). 1-Ethyl-3-(3- dimethylaminopropyl)carbodiimide hydrochloride (351 mg, 1.83 mmol) was added and the solution was stirred for 15 min. (R)-N-Methyl-2-methylamino-3-phenylpropionamide (320 mg, 1.66 mmol) dissolved in methylene chloride (4 mL) and diisopropylethylamine (0.28 mL, 1.66 mmol) were added and the mixture was stirred overnight. Methylene chloride (50 mL) was added and the organic phase was washed with water (100 mL), sodium hydrogensulfate (50 mL, 5%) and sodium hydrogen carbonate (50 mL, saturated). The organic phase was dried (magnesium sulfate) and the solvent removed in vacuo. The residue was chromatographed (silica, 2 x 45 cm) using ethylacetate/methylene chloride (1:1) to afford 604 mg of N-methyl-N- {(1 R)-1 -(N-methyl-N-((1 R)-1 -(methylcarbamoyl)-2-phenylethyl)carbamoyl)-2-(2-naphthyl)- ethyljcarbamic acid tert-butyl ester. Η-NMR (CDCI3): d 1.05, 1.31, 1.56 (three s, 9H); 2.28-3.37 (several m, 13 H); 5.04, 5.17, 5.29, 5.48 (four dd, 2H); 7.05-7.79 (m, 12 H).
(2R)-N-Methyl-2-methylamino-N-((1R)-1-(methylcarbamoyl)-2-phenylethyl)-3-(2- naphthyl)propionamide:
Figure imgf000025_0001
Sj£p ± N-Methyl-N-{(1R)-1-(N-methyl-N-((1R)-1-(methylcarbamoyl)-2-phenylethyl)carbamoyl)- 2-(2-naphthyl)ethyl}carbamic acid tert-butyl ester (600 mg, 1.19 mmol) was stirred in trifluoroacetic acid/methylene chloride (1:1, 5 mL) for 10 min and the volatiles were removed in vacuo. The residue was stripped with diethylether (2 x 5 mL) and dissolved in methanol (2 mL) and mixed with sodium hydrogen carbonate (10 mL) and ethylacetate (15 mL). The organic phase was separated and dried (magnesium sulfate) to afford 420 mg of (2R)-N-methyl-2- methylamino-N-((1R)-1-(methylcarbamoyl)-2-phenylethyl)-3-(2-naphthyl)propionamide.
Η-NMR (CDCI3): (selected values) d 1.69 (s, 3H); 2.08 (d, 3H); 2.54 (s, 3H); 2.76 (dd, 1H); 2.92 (dd, 1H), 3.12 (dd, 1H), 3.31 (dd, 1H); 3.72 (dd, 1H), 4.95 (q (br), 1H); 5.50 (dd, 1H).
((3E)-1 , 1 -Dimethyl-4-(N-methyl-N-((1 R)-1 -(N-methyl-N-((1 R)-1 -(methylcarbamoyl)-2- phenylethyl)carbamoyl)-2-(2-naphthyl)ethyl)carbamoyl)but-3-enyl)carhamic acid tert-butyl ester
Figure imgf000026_0001
Step ; (2E)-5-(tert-Butyloxycarbonylamino)-5-methylhex-2-enoic acid (200 mg, 0.82 mmol), 1- hydroxy-7-azabenzotriazole (112 mg, 0.82 mmol) and 1-ethyl-3-(3- imethylaminopropyl)- carbodiimide hydrochloride (173 mg, 0.90 mmol) were dissolved in a mixture of methylene chloride (10 mL) and N,N-dimethylformamide (1 mL) and strirred for 15 min. N-Methyl-2- methylamino-N-((1 R)-1 -(methylcarbamoyl)-2-phenylethyl)-3-(2-naphthyl)propionamide (332 mg, 0.82 mol) dissolved in methylene chloride (5 mL) and diisopropylethylamine (0.14 mL) were added and the mixture was stirred overnight under nitrogen atmosphere. The mixture was diluted with methylene chloride (50 mL), washed with water (50 mL), sodium hydrogen carbonate (30 mL, saturated), and sodium hydrogensulfate (30 mL, 5%). The phases were separated and the organic phase was dried with magnesium sulfate and evaporated in vacuo. The residue was chromatographed (silica, 2 x 40 cm) to afford 450 mg of ((3E)-1,1-dimethyl-4- (N-methyl-N-((1 R)-1 -(N-methyl-N-((1 R)-1-(methylcarbamoyl)-2-phenylethyl)carbamoyl)-2-(2- naphthyl)ethyl)carbamoyl)but-3-enyl)-carbamic acid tert-butyl ester.
Η-NMR (CDCIJ): (selected values) d 1.20, 1.22, 1.24, 1.30, 1.41, 1.55 (six s, 15 H), 4.30, 4.40 (two s (br), 1H); 5.08, 5.18, 5.32, 5.60, 5.87 (five dd, 2H); 6.05 (dd, 1H); 6.75 (m, 1H).
Stfip_Jι ((3E)-1 ,1-DimethyM-(methyl-((1R)-1-(methyl-((1R)-1-(methylcarbamoyl)-2-phenyμ ethyl)-carbamoyl)-2-(2-naphthyl)ethyl)carbamoyl)but-3-enyl)carbamic acid tert-butyl ester (403 mg, 0.63 mmol) was stirred in a mixture of trifiuroacetic acdi (4mL) and methylene chloride (4 mL) for 10 min. The volatiles were removed in vacuo and the crude product was chromatograped on silica (400g) using a mixture of methylene chloride, ethanol and ammonia (25% in water) (80/18/2) as eluent. The isolated product was dissolved in 3M hydrochloric acid in ethyl acetate and evaporated, then redissolved in methylene chloride and evaporated twice to afford 140 mg of the title compound. Η-NMR (CDCy- d 1 05, 1 10, 1 15, 1 16 (four s, 6H); 2.07 (s (br), 3H), 5.12, 5.32, 540, 5.60, 5.91 (five dd, 2H); 6.05, 6 14 (two d, 1H); 6 80 (m, 1H)
HPLC Rt = 29.02 mm (Method A1)
ESMS m/z = 529 (100%)(M+H)+
EXAMPLE 4
(2E)-5-Amino-5-methylhex-2-enoic acid N-((1R)-1-(((1R)-1-((2S)-2-hydroxypropylcarbamoyl)-2- phenylethyl)-methylcarbamoyl)-2-(2-naphthyl)ethyl)-N-methylamιde-
Figure imgf000027_0001
This compound was prepared analogously to example 1 (S)-2-hydroxypropylamιne was substituted for methylamine in step E
Η-NMR (CDCI3) (selected peaks, mixture of rotamers) d 3.90 (m, 1H); 5.55 (dd, 1H); 5.58 (d, 1H)
HPLC: R,= 29.03 mm (Method A1 )
PDMS: m/z = 573.5 (100%)(M+H)+ EXAMPLE 5:
(2E)-5-Amιno-5-methylhex-2-enoic acid ((1R)-1-(((1R)-2-(4-fluorophenyl)-1-(methylcarbamoyl)- ethyl)methylcarbamoyl)-2-(2-naphthyl)ethyl)methylamide:
Figure imgf000028_0001
(R)-2-(N-tert-Butoxycarbonyl-N-methylamino)-3-(4-fluorophenyl)propionic acid:
Figure imgf000028_0002
2-tert-Butoxycarbonylamino-3-(4-fluorophenyl)propionic acid (5.0 g; 17.7 mmol) was dissolved in dry tetrahydrofuran. lodomethane (8.8 mL; 141 mmol) was added and the reaction mixture was cooled to 0° C. Sodium hydride (2.1 g; 53.0 mmol) was slowly added and the reaction mixture was stirred for 12 hours at room temperature. Ethyl acetate (50 mL) was added and water (20 mL) was added dropwise to the reaction mixture. The ethyl acetate was removed in vacuo and the residue was diluted with diethyl ether (30 mL) and water (100 mL). The organic phase was extracted with a saturated aqueous solution of sodium hydrogen carbonate ( 50 mL). Citric acid (5 %) was added to the combined aqueous phases until pH 3, which were then extracted with ethyl acetate (2 x 50 mL) and the phases were separated. The organic phase was washed with water (2 x 50 mL), an aqueous solution of sodium thiosulfate (5 %; 2 x 50 mL) and water (50 mL) and dried (magnesium sulfate). The solvent was removed in vacuo and the residue was dissolved in diethyl ether (10 mL). Dicyclohexylamine (10 mL) was added. Methylene chloride (30 L) was added and the mixture was heated until the precipitate was dissolved Diethyl ether (20 mL) and heptane (20 L) were added and the reaction mixture was left 12 hours without stirring The reaction mixture was filtered to afford 5 7 g of (R)-2-(N- tert-butoxycarbonyl-N-methylamιno)-3-(4-fluorophenyl)propιonιc acid as a dicyclohexyl- ammonium salt
Η-NMR (CDCI3) (mixture of rotamers) d 1.21 , 1 31 (two s, 9H), 2 75, 2 84 (two s, 3H), 2.86- 3 02 (m, 1H), 3 28-3 42 (m, 1H), 4 65, 4 85 (two dd, 1H), 6 85-7 00 (m, 2H), 7 10-7.25 (m, 2H)
((1R)-2-(4-Fluorophenyl)-1-(methylcarbamoyl)ethyl)-methylcarbamιc acid tert-butylester
Figure imgf000029_0001
The dicyclohexylammoniumsalt of (R)-2-(N-tert-butoxycarbonyl-N-methylamιno)-3-(4-fluoro- phenyOpropionic acid (3 00 g, 10 1 mmol) was dissolved in methylene chloride (30 mL) and washed with an aqueous solution of sodium hydrogen sulfate (10 %, 30 mL) The organic phase was dried (magnesium sulfate) and filtered 1-Hydroxybenzotπazole (1 40 g, 10 1 mmol) and N-(3-dιmethylamιnopropyl)-N'-ethylcarbodιιmιde hydrochloride (2 0 g, 106 mmol) were added to the filtrate and the reaction mixture was stirred for 15 mm at room temperature Methylamine (40 % in methanol, 0 75 g, 9 17 mmol) and diisopropylethylamme (1 7 mL, 10 1 mmol) were added and the reaction mixture was stirred for 12 hours at room temperature The reaction mixture was washed with an aqueous solution of sodium hydrogen carbonate (sat, 50 mL) and an aqueous solution of sodium hydrogen sulfate (10 %, 50 mL) and dried (magnesium sulfate) The solvent was removed in vacuo and the residue was chromatographed on silica (3 x 40 cm) using ethyl acetate/heptane (2 1) as eluent to afford 1 06 g of ((1R)-2-(4- fluorophenyl)-1-(methylcarbamoyl)ethyl)-methylcarbamιc acid tert-butylester
1H-NMR (CDCI3) d 1 29, 1 37 (two s, 9H), 2 74 (s, 3H), 2 8 (s, 3H), 2 82-2 95 (m, 1H), 3 36- 3 48 (m, 1H), 4 63, 4 86 (m, 1H), 5 89, 6 14 (two s, 1H), 6 9-7 0 (m, 2H), 7 1-7 21 (m, 2H) (2R)-3-(4-Fluorophenyl)-N-methyl-2-(methylamino)propion-amide:
Figure imgf000030_0001
((1R)-2-(4-Fluorophenyl)-1-(methylcarbamoyl)ethyl)-methylcarbamic acid tert-butylester (1.0 g; 3.22 mmol) was dissolved in methylene chloride (5 mL). Trifluoroacetic acid (5 mL) was added and the reaction mixture was stirred for 30 min at room temperature. Methylene chloride (30 mL), an aqueous solution of sodium hydrogen carbonate/sodium carbonate (pH 9; 30 mL) and sodium hydrogen carbonate (solid), were added to the reaction mixture, until pH 9. The organic phase was dried (magnesium sulfate) and evaporated in vacuo to afford 0.62 g of (2R)-3-(4- fluorophenyl)-N-methyl-2-methylaminopropionamide.
1H-NMR (CDCI3) d : 1.31 (s, 1H); 2.29 (s, 3H); 2.65-2.73 (m, 1H); 2.82 (d, 3H); 3.12-3.20 (m, 2H); 6.96-7.02 (m, 2H); 7.11 (s, 1 H); 7.14-7.20 (m, 2H).
((1R)-1-(((1 R)-2-(4-Fluorophenyl)-1-(methylcarbamoyl)ethyl)methylcarbamoyl)-2-(2-naphthyl)- ethyl)methylcarbamic acid tert-butylester:
Figure imgf000030_0002
(2R)-2-(tert-Butoxycarbonylmethylamino)-3-(2-naphthyl)propionic acid (1.0 g; 3.1 mmol) was dissolved in methylene chloride (20 mL). 1-Hydroxy-7-azabenzotriazole (0.43 g; 3.1 mmol) and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (0.63 g; 3.3 mmol) were added and the reaction mixture was stirred for 15 min at room temperature. (2R)-3-(4-Fluorophenyl)-N-methyl-2-(methylamino)propionamide (0.6 g; 2.9 mmol) and diisopropylethylamine (0.54 mL; 3.1 mmol) was added and the reaction mixture was stirred for 12 hours at room temperature. Methylene chloride (30 mL) was added and the reaction mixture was washed with water (30 mL), an aqueous solution of sodium hydrogen sulfate (10 %; 30 mL), an aqueous solution of sodium hydrogen carbonate/sodium carbonate (pH 9; 30 mL) and water (30 mL) and dried (magnesium sulfate). The solvent was removed in vacuo and the residue was chromatographed on silica (4.0 x 30 cm) using ethyl acetate/heptane (2:1) as eluent to afford 1.07 g of ((1R)-1-(((1R)-2-(4-fluorophenyl)-1-(methylcarbamoyl)- ethyl)methylcarbamoyl)-2-(2-naphthyl)ethyl)methylcarbamic acid tert-butylester.
Η-NMR (CDCy (selected peaks for major rota er) d : 1.34 (s, 9H); 2.23 (d, 3H); 2.76 (s, 3H); 2.87 (s, 3H); 5.70 (dd, 1H); 5.95 (dd, 1H).
(2R)-N-((1R)-2-(4-Fluorophenyl)-1-(methylcarbamoyl)-ethyl)-N-methyl-2-methylamino-3-(2- naphthyhpropionamide:
Figure imgf000031_0001
((1R)-1-(((1 R)-2-(4-Fluorophenyl)-1-(methylcarbamoyl)-ethyl)methylcarbamoyl)-2-(2- naphthyl)ethyl)methylcarbamic acid tert-butylester. (1.0 g; 1.92 mmol) was dissolved in methylene chloride (5 mL). Trifluoroacetic acid (5 mL) was added and the reaction mixture was stirred for 15 min at room temperature. Methylene chloride (25 mL), an aqueous solution of sodium hydrogen carbonate/sodium carbonate (pH 9; 25 mL) and sodium hydrogen carbonate (solid) was added to the reaction mixture until pH 8. The organic phase was dried (magnesium sulfate) and evaporated in vacuo to afford 0.75 g of (2R)-N-((1R)-2-(4-fluorophenyl)-1- methylcarbamoylethyl)-N-methyl-2-methylamino-3-(2-naphthyl)propionamide. Η-NMR (CDCy d • 1 81 (s, 3H), 2.07 (d, 3H), 2.54 (s, 3H); 2.68-2.77 (m, 1 H), 2.88-2.97 (m, 1H), 3.18 (dd, 1H); 3.27 (dd, 1H), 3.8 (dd, 1H), 4 95 (s, 1H), 5 43 (dd, 1H); 6.72 (t, 1H); 6.90 (t, 2H), 7 12 (dd, 2H), 7 32 (d, 1H), 7 42-7 50 (m, 2H), 7 62 (s, 1H), 7.70-7 83 (m, 2H)
(4(((1R)-1(((1R)-2(4-Fluorophenyl)-1-(methylcarbamoyl)-ethyl)methylcarbamoyl)-2-(2- naphthyl)ethyl)methylcarbamoyl)-1 ,1-dιmethylbut-3-enyl)carbamιc acid tert-butylester
Figure imgf000032_0001
(2E)-5-(tert-Butyloxycarbonylamιno)-5-methylhex-2-enoιc acid (0.22 g, 0 89 mmol, prepared as in example 1) was dissolved in methylene chloride (10 mL) 1-Hydroxy-7-azabenzotπazole (0 13 g, 0 98 mmol) and N-(3-dιmethylamιnopropyl)-N'-ethylcarbodιιmιde hydrochloπde (0.2 g, 1.02 mmol) were added and the reaction mixture was stirred for 15 mm at room temperature (2R)-N-((1R)-2-(4-Fluorophenyl)-1-(methylcarbamoyl)ethyl)-N-methyl-2-methylamιno-3-(2- naphthyl)propιonamιde (0.38 g, 0.89 mmol) and diisopropylethylamine (0.17 mL; 0.98 mmol) were added and the reaction mixture was stirred for 12 hours at room temperature.
Methylene chlonde (50 mL) was added and the reaction mixture was washed with water (50 mL), an aqueous solution of sodium hydrogen sulfate (10 %, 50 mL), an aqueous solution of sodium hydrogen carbonate/sodium carbonate (pH 9; 50 mL) and water (50 mL) and dπed (magnesium sulfate). The solvent was removed in vacuo and the residue was chromatographed on silica (4 x 30 cm) using ethyl acetate/heptane (2:1) as eluent to afford 0.34 g of (4-(((1R)-1-(((1R)-2-(4-fluorophenyl)-1-methylcarbamoylethyl)methylcarbamoyl)-2-(2- naphthyl)-ethyl)methylcarbamoyl)-1 ,1-dιmethylbut-3-enyl)carbamιc acid tert-butyl ester
1H-NMR (CDCI3) (selected peaks for major rotamer) d . 0.85 (s, 3H), 0 87 (s, 3H), 1 42 (s, 9H), 2 12 (d, 3H), 2 72 (s, 3H), 2.96 (s, 3H), 5.75 (dd, 1H), 5,92 (dd, 1H), 6 12 (dd, 1H)
(4-(((1R)-1-(((1R)-2-(4-Fluorophenyl)-1-methylcarbamoyl-ethyl)methylcarbamoyl)-2-(2- naphthyl)ethyl)methyl-carbamoyl)-1 ,1-dιmethylbut-3-enyl)carbamιc acid tert-butylester (0 33 g, 0.51 mmol) was dissolved in methylene chloride (3 mL). Trifluoroacetic acid (3 mL) was added and the reaction mixture was stirred for 5 min at room temperature. Methylene chloride (25 L), an aqueous solution of sodium hydrogen carbonate/sodium carbonate (pH 9; 25 mL) and sodium hydrogen carbonate (solid) were added to the reaction mixture until pH 9. The organic phase was dried (magnesium sulfate) and evaporated in vacuo to afford 0.18 g of the title compound.
Η-NMR (CDCI3) (selected peaks for major rotamer) d : 1.15 (s, 6H); 2.14 (d, 3H); 2.73 (s, 3H); 3.09 (s, 3H); 5.23 (dd, 1 H); 5.90 (dd, 1H); 6.12 (dd, 1 H).
PDMS: m/z 547.4 (M+H)*
HPLC: Rt = 32.05 min

Claims

1. A method of preventing or treating osteoporosis and related disorders, the method comprising cyclic administration, to an animal in need thereof, of a growth hormone component such as a growth hormone, a somatostatin antagonist or a growth hormone secretagogue in an amount sufficient to substantially prevent or reduce the degree of osteoporosis and/or to substantially increase bone strength.
2. A method according to claim 1 , wherein the growth hormone component is administered for a period of from about 2 days to about 28 days at intervals from the termination of one period to the start of the next period of from about 1 week to about 26 weeks.
3. A method according to claim 2, wherein the growth hormone component is administered for a period of about 7 days at intervals from the start of one period to the start of the next period of about 6 weeks to about 12 weeks.
4. A method according to claim 1, wherein the animal is a mammal, in particular a human being.
5. A method according to claim 1 , wherein the growth hormone component is human growth hormone.
6. A method according to claim 1 , wherein a composition with anti-resoφtive action on bone is continuously administered to the animal in addition to the cyclic administration of the growth hormone component.
7. A method according to claim 6, wherein said composition comprises an estrogen, in particular estradiol.
8. A method according to claim 6, wherein said composition comprises a compound with estrogenic effect.
9. A method according to claim 6, wherein said composition comprises one or more compounds selected from the group comprising Centchroman, Levormeloxifene, Raloxifene, Droloxifene, Tamoxifene, or Idoxifene.
10. A method according to claim 6, wherein said composition comprises a calcitonin.
11. A method according to claim 6, wherein said composition comprises a bisphosphonate.
12. A method according to any of the claims 7 to 11 , wherein the composition comprises a combination of an estrogen and a gestagen.
13. A method according to any one of the preceding claims, wherein the dose or release of growth hormone is in the range of about 0.01-1 lU/kg body weight/day, in particular about 0.2 lU/kg body weight/day.
14. A method according to claim 6, wherein the dose of the composition with an anti-resoφtive action on bone is in the range of from 0.001 to 10 mg/kg body weight/day and wherein the dose of growth hormone is in the range of about 0.01-1 lU/kg body weight/day, in particular about 0.2 lU/kg body weight/day.
15. Use of a growth hormone component for the manufacture of a medicament for the prevention or treatment of osteoporosis and related diseases, the medicament being intended for cyclic administration.
16. Use of a growth hormone component for the manufacture of a medicament for the prevention or treatment of osteoporosis and related diseases, the medicament being packaged with instructions for cyclic administration thereof.
17. Use of a growth hormone or a growth hormone component together with a composition with an anti-resoφtive action on bone for the manufacture of a medicament for the prevention or treatment of osteoporosis and related diseases, the medicament being intended for cyclic administration of the growth hormone component and continuous administration of the anti- resorptive composition. 18 Use of a growth hormone component together with a composition with an anti-resoφtive action on bone for the manufacture of a medicament for the prevention or treatment of osteoporosis and related diseases, the medicament being packaged with instructions for cyclic administration of the growth hormone component and continuous administration of the anti- resorptive composition
19 Use of a growth hormone for the manufacture of a medicament for the prevention or treatment of osteoporosis and related diseases, the medicament being packaged with instructions for cyclic administration of the growth hormone component together with continuous administration of the anti-resorptive composition
20 Use of a composition with an anti-resorptive action on bone for the manufacture of a medicament for the prevention or treatment of osteoporosis and related diseases, the medicament being intended for continuous administration over a period of time concomitantly with cyclic administration of a growth hormone component
21 Products containing a) a growth hormone component and b) a compound with anti- resoφtive effect on bone as a combined preparation for simultaneous, separate or sequential use in the prevention or treatment of osteoporosis
22 A method according to claim 9, wherein said composition compπses Levormeloxifene or a structurally related compound thereof
23 A method according to claim 9, wherein said composition comprises Levormeloxifene
PCT/DK1997/000239 1996-05-31 1997-05-29 Growth hormone component and bone anti-resorptive agent in cyclic (coherence) treatment of osteoporosis WO1997046252A1 (en)

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Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB2324726A (en) * 1997-05-01 1998-11-04 Merck & Co Inc Combination Therapy for the Treatment of Osteoporosis
WO1999064456A1 (en) * 1998-06-09 1999-12-16 Novo Nordisk A/S A method for preparing a compound with growth hormone releasing properties
EP1069109A1 (en) * 1999-07-14 2001-01-17 Ajinomoto Co., Inc. Process for production of optically active N-protected-N-methyl-phenylalanine derivative
EP1159964A2 (en) 2000-05-31 2001-12-05 Pfizer Products Inc. Compositions and methods for stimulating gastrointestinal motility
US6784158B1 (en) 1998-06-09 2004-08-31 Novo Nordisk A/S Method for preparing a compound with growth hormone releasing properties
US7255984B2 (en) 1996-02-28 2007-08-14 Pfizer Inc. Combination therapy for osteoporosis
WO2007098716A1 (en) 2006-02-28 2007-09-07 Centro De Ingeniería Genética Y Biotecnología Compounds analogous to growth hormone peptide secretagogues and preparations containing them
EP2457893A1 (en) 2004-06-18 2012-05-30 Tranzyme Pharma, Inc. Intermediates for macrocyclic modulators of the ghrelin receptor
EP2644618A1 (en) 2007-02-09 2013-10-02 Tranzyme Pharma, Inc. tether intermediates for the synthesis of macrocyclic ghrelin receptor modulators

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0436469A1 (en) * 1990-01-03 1991-07-10 Ciba-Geigy Ag Composition and method for the treatment of osteoporosis in mammals
WO1992009304A1 (en) * 1990-11-26 1992-06-11 Recker Robert R Treatment for osteoporosis using growth hormone release factor (grf) in combination with parathyroid hormone (pth)
WO1995011029A1 (en) * 1993-10-19 1995-04-27 Merck & Co., Inc. Combination of bisphosphonates and growth hormone secretagogues

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0436469A1 (en) * 1990-01-03 1991-07-10 Ciba-Geigy Ag Composition and method for the treatment of osteoporosis in mammals
WO1992009304A1 (en) * 1990-11-26 1992-06-11 Recker Robert R Treatment for osteoporosis using growth hormone release factor (grf) in combination with parathyroid hormone (pth)
WO1995011029A1 (en) * 1993-10-19 1995-04-27 Merck & Co., Inc. Combination of bisphosphonates and growth hormone secretagogues

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
CALCIF TISSUE INT., Volume 40, 1987, JOHN F. ALOIA et al., "Coherence Treatment of Postmenopausal Osteoporosis with Growth Hormone and Calcitonin", pages 253-259. *
CLINICAL ENDOCRINOLOGY, Volume 46, 1997, STEFANO GONNELLI et al., "Treatment of Post-Menopausal Osteoporosis with Recombinant Human Growth Hormone and Salmon Calcitonin: a Placebo Controlled Study", pages 55-61. *
FILE 73:EMBASE, 9775990, ERDTSIECK R.J. et al., "Treatment of Post-Menopausal Osteoporosis with a Combination of Growth Hormone and Pamidronate: a Placebo Controlled Trial"; & CLINICAL ENDOCRINOLOGY, 1995, 43/5, (557-565). *
JOURNAL OF CLINICAL ENDOCRINOLOGY AND METABOLISM, Volume 82, No. 4, 1997, LEAH HOLLOWAY et al., "Skeletal Effects of Cyclic Recombinant Human Growth Hormone and Salmon Calcitonin in Osteopenic Postmenopausal Women", pages 1111-1117. *

Cited By (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7255984B2 (en) 1996-02-28 2007-08-14 Pfizer Inc. Combination therapy for osteoporosis
US6043026A (en) * 1997-05-01 2000-03-28 Merck & Co., Inc. Combination therapy for the prevention and treatment of osteoporosis
GB2324726A (en) * 1997-05-01 1998-11-04 Merck & Co Inc Combination Therapy for the Treatment of Osteoporosis
JP2002517514A (en) * 1998-06-09 2002-06-18 ノボ ノルディスク アクティーゼルスカブ Method for preparing compounds with growth hormone releasing properties
WO1999064456A1 (en) * 1998-06-09 1999-12-16 Novo Nordisk A/S A method for preparing a compound with growth hormone releasing properties
US6784158B1 (en) 1998-06-09 2004-08-31 Novo Nordisk A/S Method for preparing a compound with growth hormone releasing properties
EP1069109A1 (en) * 1999-07-14 2001-01-17 Ajinomoto Co., Inc. Process for production of optically active N-protected-N-methyl-phenylalanine derivative
US6342629B1 (en) 1999-07-14 2002-01-29 Ajinomoto Co., Inc. Process for production of optically active n-protected-n-methyl-phenylalanine derivative
EP1159964A2 (en) 2000-05-31 2001-12-05 Pfizer Products Inc. Compositions and methods for stimulating gastrointestinal motility
EP2457893A1 (en) 2004-06-18 2012-05-30 Tranzyme Pharma, Inc. Intermediates for macrocyclic modulators of the ghrelin receptor
EP2457925A1 (en) 2004-06-18 2012-05-30 Tranzyme Pharma, Inc. Process for preparing a macrocyclic modulator of the ghrelin receptor and intermediates
WO2007098716A1 (en) 2006-02-28 2007-09-07 Centro De Ingeniería Genética Y Biotecnología Compounds analogous to growth hormone peptide secretagogues and preparations containing them
EP2644618A1 (en) 2007-02-09 2013-10-02 Tranzyme Pharma, Inc. tether intermediates for the synthesis of macrocyclic ghrelin receptor modulators

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