WO1999066945A1 - Compositions and methods for inhibiting bone resorption - Google Patents
Compositions and methods for inhibiting bone resorption Download PDFInfo
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- WO1999066945A1 WO1999066945A1 PCT/US1999/013961 US9913961W WO9966945A1 WO 1999066945 A1 WO1999066945 A1 WO 1999066945A1 US 9913961 W US9913961 W US 9913961W WO 9966945 A1 WO9966945 A1 WO 9966945A1
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
- A61K38/07—Tetrapeptides
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
- A61K38/06—Tripeptides
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- 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
- A61K38/55—Protease inhibitors
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- 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
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- 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
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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
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/08—Tripeptides
- C07K5/0827—Tripeptides containing heteroatoms different from O, S, or N
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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
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/10—Tetrapeptides
- C07K5/1027—Tetrapeptides containing heteroatoms different from O, S, or N
Definitions
- compositions useful herein comprise the combination of a pharmaceutically effective amount of a nitrogen-containing bisphosphonate or a pharmaceutically-acceptable salt thereof and a pharmaceutically effective amount of a caspase inhibitor.
- disorders in humans and other mammals involve or are associated with abnormal bone reso ⁇ tion.
- Such disorders include, but are not limited to, osteoporosis, Paget's disease, periprosthetic bone loss or osteolysis, and hypercalcemia of malignancy.
- the most common of these disorders is osteoporosis, which in its most frequent manifestation occurs in postmenopausal women.
- Osteoporosis is a systemic skeletal disease characterized by a low bone mass and microarchitectural deterioration of bone tissue, with a consequent increase in bone fragility and susceptibility to fracture. Osteoporotic fractures are a major cause of morbidity and mortality in the elderly population.
- osteoporotic fracture As many as 70% of women and a third of men will experience an osteoporotic fracture. A large segment of the older population already has low bone density and a high risk of fractures. There is a significant need to both prevent and treat osteoporosis and other conditions associated with bone resorption. Because osteoporosis, as well as other disorders associated with bone loss, are generally chronic conditions, it is believed that appropriate therapy will typically require chronic treatment.
- Multinucleated cells called osteoclasts are responsible for causing bone loss through a process known as bone resorption. It is well known that bisphosphonates are selective inhibitors of osteoclastic bone resorption, making these compounds important therapeutic agents in the treatment or prevention of a variety of generalized or localized bone disorders caused by or associated with abnormal bone resorption. See H. Fleisch, Bisphosphonates In Bone Disease, From The Laboratory To The Patient, 3rd Edition, Parthenon Publishing (1997), which is incorporated by reference herein in its entirety. Without being limited by theory, it is believed that bisphophonates inhibit osteoclast function by triggering apoptosis, i.e. programmed cell death. See D.E.
- the bisphosphonate, pamidronate has been associated with esophageal ulcers.
- E.G. Lufkin et al. Pamidronate: An Unrecognized Problem in Gastrointestinal Tolerability, Osteoporosis International, 4: 320-322 (1994), which is inco ⁇ orated by reference herein in its entirety.
- the use of alendronate has been associated with esophagitis and/or esophageal ulcers. See P.C. De Groen, et al., Esophagitis Associated With The Use Of Alendronate, New England Journal of Medicine, vol. 335, no. 124, pp. 1016-1021 (1996), D.O.
- Bisphosphonate treatment regimens normally involve the chronic administration of relatively low doses of the bisphosphonate compound, with the objective of delivering the desired cumulative therapeutic dose over the course of the treatment period.
- chronic dosing especially chronic daily dosing, can have the disadvantage of causing adverse gastrointestinal effects due to the repetitive, continuous, and additive irritation to the gastrointestinal tract. This potential problem can therefore interfere with patient compliance, and in severe cases even require cessation of treatment. Therefore, it is seen that there is a need to minimize the potential adverse effects that can be associated with bisphosphonate bone reso ⁇ tion therapy.
- Caspases belong to a broad class of enzymes known as proteases, which are enzymes that hydrolyze peptide bonds. Specfically, caspases are cysteine proteases that preferentially target and cleave peptide sequences having an aspartic acid moiety. Caspases are believed to be involved in the normal cell turnover process and are mediators of apoptosis. See Nicholson and Thornberry, 1997, "Caspases: killer proteases", TIBS , vol.
- caspase inhibitors block the bisphosphonate-induced osteoclast apoptosis.
- caspase inhibitors such as Z-Val- Ala-Asp-FMK, Z-Asp- Glu-Val-Asp-FMK ,and Z-Tyr-Val- Ala-Asp-FMK can block the inhibitory effect that would otherwise be observed with a bisphosphonate such as alendronate monosodium trihydrate.
- Mst mammalian Sterile-20-like kinase
- Mst 1 a kinase known as Mst 1
- Mst 2 a kinase known as Mst 1
- Taylor et al. "Newly identified stress-responsive protein kinases, Krs-1 and Krs-2", Proc. Natl. Acad. Sci. USA, Vol. 93 (1996), pp. 10099-10104
- Creasy et al "Cloning and characterization of a human protein kinase with homology to Ste20," The J. of Biological Chemistry, Vol.
- a caspase inhibitor such as Z-VAD-FMK is found to prevent the cleavage of Mst that is induced by bisphosphonates such as alendronate monosodium trihydrate.
- caspase inhibitors have not previously been investigated either in vitro or in vivo for their ability to mitigate the potentially adverse gastrointestinal side effects that can be associated with bisphosphonate anti-bone reso ⁇ tion therapy.
- the combination of a nitrogen-containing bisphosphonate or a pharmaceutically-acceptable salt thereof and a caspase inhibitor is highly effective for inhibiting bone reso ⁇ tion while mitigating the potentially adverse gastrointestinal effects that can be associated with bisphosphonate therapy.
- the combination has the advantage of providing increased safety and better patient compliance, which should maximize therapeutic efficacy. Without being limited by theory it is believed that the caspase inhibitor blocks the potentially harmful effect of the bisphosphonate on the epithelial cells of the gastrointestinal tract.
- the caspase inhibitor By selecting an appropriate dosage of the caspase inhibitor it is possible to orally deliver a sufficiently high local concentration of the caspase inhibitor to the gastrointestinal tract to block the potentially harmful effects of the nitrogen-containing bisphosphonate, while minimizing the blocking effect on the osteoclasts, where the full therapeutic benefit of the bisphosphonate is desired to inhibit bone reso ⁇ tion.
- compositions comprising the combination of a nitrogen-containing bisphosphonate or a pharmaceutically-acceptable salt thereof and a caspase inhibitor. It is another object of the present invention to provide improved oral methods for inhibiting bone reso ⁇ tion and the conditions associated therewith in a mammal, particularly wherein said mammal is a human.
- the present invention relates to a pharmaceutical composition
- a pharmaceutical composition comprising a nitrogen-containing bisphosphonate or pharmaceutically acceptable salt thereof and a caspase inhibitor.
- the present invention relates to a pharmaceutical composition
- a pharmaceutical composition comprising a pharmaceutically-effective amount of a nitrogen-containing bisphosphonate or pharmaceutically acceptable salt thereof and an amount of a caspase inhibitor effective to counteract nitrogen-containing bisphosphonate-associated gastrointestional effects.
- the present invention relates to a method for inhibiting bone reso ⁇ tion in a mammal in need thereof comprising administering a nitrogen-containing bisphosphonate or pharmaceutically acceptable salt thereof and a caspase inhibitor.
- the present invention relates to a method for inhibiting bone reso ⁇ tion in a mammal in need thereof comprising sequentially administering a caspase inhibitor and a ntirogen-containing bisphosphonate or pharmaceutically acceptable salt thereof.
- the present invention relates to the use of a composition in the manufacture of a medicament for inhibiting bone reso ⁇ tion in a mammal in need thereof, said composition comprising a nitrogen-containing bisphosphonate or pharmaceutically acceptable salt thereof and a caspase inhibitor.
- the present invention relates to the use of a composition comprising a nitrogen-containing bisphosphonate or pharmaceutically acceptable salt thereof and a caspase inhibitor for inhibiting bone reso ⁇ tion in a mammal in need thereof. All percentages and ratios used herein, unless otherwise indicated, are by weight.
- the invention hereof can comprise, consist of, or consist essentially of the essential as well as optional ingredients, components, and methods described herein.
- FIGURES Figure 1 shows the identification of a 34 kDa Mst kinase fragment that is activated by 30 ⁇ M alendronate monosodium trihydrate (a nitrogen-containing bisphosphonate).
- the 34 kDa Mst kinase fragment is an amino-terminal cleavage product of Mst.
- Osteoclast-like cells are isolated and treated with alendronate monosodium trihydrate.
- Cell lysates are exposed to treatment with a cocktail of commercially available anti-Mst and anti-Krs antibodies that recognize the amino- terminus of Mst kinase and Protein A-agarose beads to immunoprecipitate Mst from the samples.
- Osteoclast like cells are treated with alendronate monosodium trihydrate, etidronate disodium, or tiludronate disodium at concentrations of both 30 ⁇ M and 100 ⁇ M for about 17 hours.
- Kinase activities are analyzed using the in-gel kinase assays. Autoradiographs are developed after film exposure for several days. These data show the activation of the 34 kDa Mst Kinase fragment by alendronate monosodium trihydrate at either 30 ⁇ M (lane 3) or 100 ⁇ M (lane 4).
- Figure 3 shows the preferential response of 34 kDa Kinase to alendronate monosodium trihydrate and the inability of alendronate monosodium trihydrate to activate the 34 kDa Kinase when used to treat osteoclasts in the presence of the caspase inhibitor Z-Val- Ala-Asp-FMK.
- Osteoclast-like cells are treated with alendronate monosodium trihydrate at concentrations of 10 ⁇ M and 30 ⁇ M for about 17 hours.
- Kinase activities are analyzed using an in-gel kinase assay. Autoradiographs are developed after film exposure for several days.
- Figure 4 shows that activation of Mst 1 by 30 ⁇ M alendronate monosodium trihydrate is blocked by caspase inhibitors. Osteoclast-like-cells are purified from cocultures by sequential treatment of culture dishes wih collagenase and EDTA. After the desired treatment, cell lysates are analyzed by an in-gel kinase assay using myelin basic protein as a substrate. Lane 1 is a no-treatment control. Lane 2 shows treatment with 20 ⁇ M Z-Asp-Glu-Val-Asp-FMK. Lane 3 shows treatment with 20 ⁇ M Z-Val-Ala-Asp-FMK. Lane 4 shows treatment with 20 ⁇ M Z-Tyr-Val- Ala-Asp-FMK.
- Lane 5 shows treatment with 30 ⁇ M alendronate monosodium trihydrate.
- Lane 6 shows treatment with 30 ⁇ M alendronate monosodium trihydrate and 20 ⁇ M Z-Asp-Glu-Val-Asp-FMK.
- Lane 7 shows treatment with 30 ⁇ M alendronate monosodium trihydrate and 20 ⁇ M Z-Val- Ala-Asp-FMK.
- Lane 8 shows treatment with 30 ⁇ M alendronate monosodium trihydrate and 20 ⁇ M Z-Tyr-Val-Ala- Asp-FMK.
- the present invention relates to compositions and methods for inhibiting bone reso ⁇ tion in a mammal in need of such treatment, while counteracting the occurence of adverse gastrointestinal effects.
- the compositions comprise a pharmaceutically effective amount of a nitrogen-containing bisphosphonate or a pharmaceutically-acceptable salt thereof and a pharmaceutically effective amount of a caspase inhibitor.
- pharmaceutically effective amount means that amount of the nitrogen-containing bisphosphonate compound or caspase inhibitor, that will elicit the desired therapeutic effect or response or provide the desired benefit when administered in accordance with the desired treatment regimen.
- a prefered pharmaceutically effective amount of the nitrogen-containing bisphosphonate is a bone reso ⁇ tion inhibiting amount.
- a preferred pharmaceutically effective amount of the caspase inhibitor is an amount that will counteract, i.e. block or mitigate, the occurrence of adverse gastrointestinal effects, while not counteracting, or only minimally counteracting, the therapeutic bone reso ⁇ tion effects of the nitrogen-containing bisphosphonate.
- adverse gastrointestinal effects means to prevent, block, decrease, or lessen the occurrence of unwanted side effects in the gastrointestinal tract, i.e. the esophagus, stomach, intestines, and rectum, particularly the esophagus, relative to treatment with a nitrogen-containing bisphosphonate alone.
- adverse gastrointestinal effects include, but are not limited to those selected from the group consisting of esophagitis, esophageal ulcers, esophageal irritation, esophageal perforation, abdominal pain, gastric duodenal ulcers, and constipation.
- abnormal bone reso ⁇ tion means a degree of bone reso ⁇ tion that exceeds the degree of bone formation, either locally, or in the skeleton as a whole.
- abnormal bone reso ⁇ tion can be associated with the formation of bone having an abnormal structure, as in Paget's disease.
- bone reso ⁇ tion inhibiting mean's preventing bone reso ⁇ tion by the direct or indirect alteration of osteoclast formation or activity. Inhibition of bone reso ⁇ tion refers to prevention of bone loss, especially the inhibition of removal of existing bone either from the mineral phase and/or the organic matrix phase, through direct or indirect alteration of osteoclast formation or activity.
- the term "until the desired therapeutic effect is achieved”, as used herein, means that the therapeutic agent or agents are continuously administered, according to the dosing schedule chosen, up to the time that the clinical or medical effect sought for the disease or condition being treated is observed by the clinician or researcher. For methods of treatment of the present invention, the pharmaceutical composition is continuously administered until the desired change in bone mass or structure is observed.
- administration periods can range from about 2 weeks to the remaining lifespan of the mammal.
- administration periods can range from about 2 weeks to the remaining lifespan of the human, preferably from about 2 weeks to about 20 years, more preferably from about 1 month to about 20 years, more preferably from about 6 months to about 10 years, and most preferably from about 1 year to about 10 years.
- nitrogen-containing means that the bisphosphonate compound or pharmaceutically acceptable salt thereof comprises at least one nitrogen atom in the bisphosphonate portion of the molecule.
- any nitrogen atom contained in the positive counter ion of such a salt e.g., the nitrogen atom of an ammonium counter ion, would not be considered in meeting the "nitrogen-containing" definition.
- alendronic acid i.e. 4-amino-l -hydroxybutylidene- 1,1- bisphosphonic acid is an example of a nitrogen-containing bisphosphonate.
- the ammonium salt of the unsubstituted 1 -hydroxybutylidene- 1 , 1 -bisphosphonic acid would not be a nitrogen-containing bisphosphonate as defined herein.
- compositions of the present invention are provided.
- compositions of the present invention comprise a pharmaceutically effective amount of a nitrogen-containing bisphosphonate or a pharmaceutically-acceptable salt thereof and a pharmaceutically effective amount of a caspase inhibitor. These compositions are useful for inhibiting bone reso ⁇ tion in a mammal in need thereof while counteracting the potentially adverse effects, such as gastrointestinal effects, that can be associated with the administration of the bisphosphonate.
- a and X are independently selected from the group consisting of H, OH, OR, halogen, NH2, NHR, N 2, SR, SH, C 1 -C30 alkyl (including straight, branched, and cyclic alkyl), substituted C1-C30 alkyl (including straight, branched, and cyclic alkyl), and C5-C14 aryl, wherein said substituted C1-C30 alkyl comprises one or more moieties selected from the group consisting of OH, OR, halogen, NH2, NHR, NR2, SH, SR, and C5-C14 aryl, wherein R is selected from the group consisting of C1-C30 alkyl (including straight, branched, and cyclic alkyl) and C5- C14 aryl, with the proviso that the resulting bisphosphonate compound contains at least one nitrogen atom.
- a and X can be taken together to form a cyclic moiety.
- Nonlimiting examples of the substituents for the substituted C1-C30 alkyl moiety include those selected from the group consisting of amino, alkylamino, dialkylamino, imidazolyl, pyridyl, imidazonyl, imidazopyridinyl, furanyl, and cycloalkylamino.
- aryl refers to a monocyclic or polycyclic system comprising at least one aromatic ring, wherein the monocylic or polycyclic system contains 0, 1, 2, 3, or 4 heteroatoms chosen from N, O, or S, and wherein the monocyclic or polycyclic system is either unsubstituted or substituted with one or more groups independently selected from hydrogen, halogen, Ci-io alkyl, C3-8 cycloalkyl, aryl, aryl Ci-8 alkyl, amino, amino Ci-8 alkyl, C ⁇ _3 acylamino, Ci-3 acylamino Ci-8 alkyl, Ci-6 alkylamino, Ci-6 alkylamino Ci-8 alkyl, Ci-6 dialkylamino, Ci-6 dialkylamino-Ci-8 alkyl, Ci-4 alkoxy, Ci-4 alkoxy Ci-6 alkyl, hydroxycarbonyl, hydroxycarbonyl Ci-6 alkyl, Ci-5 alkoxycarbonyl, Ci-3 alk
- aryl examples include, but are not limited to, phenyl, naphthyl, pyridyl, pyrazinyl, pyrimidinyl, imidazolyl, benzimidazolyl, indolyl, thienyl, furyl, dihydrobenzofuryl, benzo(l,3) dioxolane, oxazolyl, isoxazolyl and thiazolyl.
- Preferred nitrogen-containing bisphosphonates are those in which A is selected from the group consisting of H, OH, and halogen, more preferably OH, and X is selected from the group consisting of NH2, NHR, NR2, C5-C14 aryl, substituted
- C1-C30 alkyl (including straight, branched, and cyclic alkyl) wherein said substituted alkyl group is substituted with one or more moieties selected from the group consisting of OH, OR, halogen, NH2, NHR, NR2, SH, SR, and C5-C14 aryl, with the proviso that the resulting compound contains at least one nitrogen atom.
- A is OH and X is a 3-aminopropyl moiety, so that the resulting compound is a 4-amino-l -hydroxybutylidene- 1,1 -bisphosphonate, i.e. alendronate.
- salts and derivatives of the nitrogen- containing bisphosphonates are also useful herein.
- Nonlimiting examples of salts include those selected from the group consisting of alkali metal, alkaline metal, ammonium, and mono-, di, tri-, or tetra-Cl-C30-alkyl-substituted ammonium.
- Preferred salts are those selected from the group consisting of sodium, potassium, calcium, magnesium, and ammonium salts.
- the salts are formed when one or more of the hydrogen atoms of the phosphonate groups are substituted.
- Nonlimiting examples of derivatives include those selected from the group consisting of esters and amides.
- bisphosphonate and “bisphosphonates”, as used herein in referring to the nitrogen-containing bisphosphonates are meant to also encompass the terms diphosphonates, biphosphonic acids, and diphosphonic acids, as well as salts and derivatives of these materials.
- the use of a specific nomenclature in referring to the bisphosphonate or bisphosphonates is not meant to limit the scope of the present invention, unless specifically indicated. Because of the mixed nomenclature currently in use by those or ordinary skill in the art, reference to a specific weight or percentage of a bisphosphonate compound in the present invention is on an acid active weight basis, unless indicated otherwise herein.
- the phrase "about 10 mg of a nitrogen-containing bisphosphonate selected from the group consisting of alendronate, pharmaceutically acceptable salts thereof, and mixtures thereof, on an alendronic acid active weight basis" means that the amount of the bisphosphonate compound selected is calculated based on 10 mg of alendronic acid.
- Nonlimiting examples of nitrogen-containing bisphosphonates useful herein include the following: Alendronic acid, 4-amino- 1 -hydroxybutylidene- 1 , 1 -bisphosphonic acid.
- Alendronate also known as alendronate sodium or monosodium trihydrate
- 4-amino-l -hydroxybutylidene- 1,1 -bisphosphonic acid monosodium trihydrate Alendronic acid and alendronate are described in U.S. Patents 4,922,007, to Kieczykowski et al., issued May 1, 1990, and 5,019,651, to Kieczykowski, issued May 28, 1991, both of which are inco ⁇ orated by reference herein in their entirety.
- Cycloheptylaminomethylene- 1,1 -bisphosphonic acid, YM 175 Cycloheptylaminomethylene- 1,1 -bisphosphonic acid
- [l-hydroxy-2-imidazo-(l,2-a)pyridin-3-ylethylidene]bis- phosphonate (Yamanouchi YH 529).
- alendronate More preferred is alendronate, pharmaceutically acceptable salts thereof, and mixtures thereof. Most preferred is alendronate monosodium trihydrate. It is recognized that mixtures of two or more of the nitrogen-containing bisphosphonates can be utilized.
- the precise dosage of the nitrogen-containing bisphosphonate will vary with the dosing schedule, the particular bisphosphonate chosen, the age, size, sex and condition of the mammal or human, the nature and severity of the disorder to be treated, and other relevant medical and physical factors. Thus, a precise pharmaceutically effective amount cannot be specified in advance and can be readily determined by the caregiver or clinician. Appropriate amounts can be determined by routine experimentation from animal models and human clinical studies. Generally, an appropriate amount of nitrogen-containing bisphosphonate is chosen to obtain a bone reso ⁇ tion inhibiting effect, i.e. a bone reso ⁇ tion inhibiting amount of the nitrogen-containing bisphosphonate is administered.
- an effective oral dose of nitrogen-containing bisphosphonate is typically from about 1.5 to about 6000 ⁇ g/kg body weight and preferably about 10 to about 2000 ⁇ g/kg of body weight.
- alendronate monosodium trihydrate common human doses which are administered are generally in the range of about 2 mg/day to about 40 mg/day, preferably about 5 mg/day to about 40 mg/day.
- dosages for alendronate monosodium trihydrate are 5 mg/day for preventing osteoporosis, 10 mg/day for treating osteoporosis, and 40 mg/day for treating Paget's disease.
- the nitrogen-containing bisphosphonate can be administered at intervals other than daily, for example once-weekly dosing, twice-weekly dosing, biweekly dosing, and twice-monthly dosing.
- appropriate multiples of the bisphosphonate dosage would be administered.
- alendronate monosodium trihydrate would be administered at dosages of 35 mg/week or 70 mg/week in lieu of seven consecutive daily dosages of 5 mg or 10 mg.
- compositions herein comprise from about 1 mg to about 100 mg of nitrogen-containing bisphosphonate, preferably from about 2 mg to 70 mg, and more preferably from about 5 mg to about 70, on a bisphosphonic acid basis.
- the pharmaceutical compositions useful herein comprise about 2.5 mg, 5 mg, 10 mg, 35, mg, 40 mg, or 70 mg of the active on an alendronic acid active weight basis.
- compositions of the present invention comprise a pharmaceutically effective amount of a caspase inhibitor.
- the caspase inhibitors useful herein are generally relatively short aspartic acid-containing peptides, although non-peptide inhibitors are also intended as being within the scope of the present invention.
- “relatively short”, as used herein means that the peptides typically contain from about 3 to about 5 amino acids in length.
- “aspartic acid-containing” is meant that these peptides comprise at least one aspartic acid moiety, preferably at the carboxy-terminal end. These peptides are preferably blocked at both the amino and carboxy terminal ends with blocking groups.
- the caspase inhibitors useful herein can be represented by the following chemical formula
- amino acid (AA) can be selected from any of the naturally occurring amino acids, the D-enantiomers of the naturally-occurring amino acids (for example, D-alanine), and non-naturally occurring amino acids (for example, 3-aminopropionic acid and N-methyl glycine).
- (ATBG) and (CTBG) moities are selected from any of the blocking groups that are well known to peptide chemists of ordinary skill in the art. See Greene, T.W. et al., Protecting Groups in Organic Synthesis, 2nd edition, 1991, John Wiley & Sons, Inc., which is inco ⁇ orated by reference herein in its entirey.
- Nonlimiting examples of (ATBG) moieties are bezyloxycarbonyl group (also known as the cbz or Z group) and the t-butoxycarbonyl group (also known as the boc group), and the acyl group.
- Nonlimiting examples of (CTBG) moieties are alkyl groups (for example methyl and ethyl esters), the benzyl group, and the fluoromethyl keto group [which is abbreviated as (OMe)-CH2F or FMK].
- CTBG chloride-based compound
- OMe fluoromethyl keto group
- caspase inhibitors useful herein are disclosed in U.S. Patent No. 5,210, 272, to Palmer, issued May 11, 1993, U.S. Patent 5,101,068, to Palmer et al., issued March 31, 1992, and U.S. Patent No. 4,518,528, to Rasnick, issued May 21, 1985, which are all inco ⁇ orated by reference herein in their entirety.
- Preferred caspase inhibitors useful herein are selected from the group consisting of Z- Val-Ala-Asp-FMK (which has a molecular weight of about 468), Z-Asp-Glu-Val- Asp-FMK (which has a molecular weight of about 668), and Z-Tyr-Val- Ala-Asp- FMK (which has a molecular weight of about 630) MW 630.
- Z- Val-Ala-Asp-FMK which has a molecular weight of about 468
- Z-Asp-Glu-Val- Asp-FMK which has a molecular weight of about 668
- Z-Tyr-Val- Ala-Asp- FMK which has a molecular weight of about 630
- the precise dosage of the caspase inhibitor will vary with the dosing schedule, the particular compound chosen, the age, size, sex and condition of the mammal or human, the nature and severity of the disorder to be treated, and other relevant medical and physical factors. Thus, a precise pharmaceutically effective amount cannot be specified in advance and can be readily determined by the caregiver or clinician. Appropriate amounts can be determined by routine experimentation from animal models and human clinical studies. Generally, an appropriate amount is chosen to obtain an inhibition of the potentially adverse gastrointestinal effects of the nitrogen-containing bisphosphonate. The amount should be below that level which will inhibit the desired bone reso ⁇ tion inhibiting effect of the nitrogen-contianing bisphosphonate.
- an effective oral dose of the caspase inhibitor is typically chosen so as to provide a local concentration in the esophagus from about 1 ⁇ M to about 100 ⁇ M, preferably about 10 ⁇ M, although other ranges can be used.
- Nonlimiting exemplary doses are about 1 ug/kg to about 100 ug/kg, preferably about 10 ug/kg, for a human subject.
- human doses which can be administered are generally in the range of about 0.1 mg/day to about 10 mg/day, preferably from about 0.25 mg/day to about 5 mg/day, and more preferably from about 0.5 mg/day to about 1.5 mg/day.
- a typical nonlimiting dosage amount would be about 0.75 mg/day.
- the pharmaceutical compositions herein comprise from about 0.1 mg to about 10 mg, preferably from about 0.25 mg to about 5 mg, and more preferably from about 0.5 mg to about 1.5 mg of the caspase inhibitor.
- a typical nonlimiting amount for is about 0.75 mg.
- the nitrogen-containing bisphosphonate and the caspase inhibitor are typically administered in admixture with suitable pharmaceutical diluents, excipients, or carriers, collectively referred to herein as "carrier materials”, suitably selected with respect to oral administration, i.e. tablets, capsules, elixirs, syrups, powders, and the like, and consistent with conventional pharmaceutical practices.
- carrier materials suitably selected with respect to oral administration, i.e. tablets, capsules, elixirs, syrups, powders, and the like, and consistent with conventional pharmaceutical practices.
- the active ingredient can be combined with an oral, non-toxic, pharmaceutically acceptable inert carrier such as lactose, starch, sucrose, glucose, methyl cellulose, magnesium stearate, mannitol, sorbitol, croscarmellose sodium and the like;
- an oral, non-toxic, pharmaceutically acceptable inert carrier such as lactose, starch, sucrose, glucose, methyl cellulose, magnesium stearate, mannitol, sorbitol, croscarmellose sodium and the like
- the oral drug components can be combined with any oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like.
- suitable binders, lubricants, disintegrating agents and coloring agents can also be inco ⁇ orated.
- Suitable binders can include starch, gelatin, natural sugars such a glucose, anhydrous lactose, free-flow lactose, beta-lactose, and corn sweeteners, natural and synthetic gums, such as acacia, guar, tragacanth or sodium alginate, carboxymethyl cellulose, polyethylene glycol, waxes, and the like.
- Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like.
- a particularly preferred tablet formulation for alendronate monosodium trihydrate is that described in U.S. Patent No.
- the compounds used in the present method can also be coupled with soluble polymers as targetable drug carriers.
- soluble polymers can include polyvinylpyrrolidone, pyran copolymer, polyhydroxylpropyl-methacrylamide, and the like.
- the present invention comprises methods for treating abnormal bone reso ⁇ tion in mammals.
- the present invention also comprises methods for preventing abnormal bone reso ⁇ tion in mammals.
- the mammal is a human.
- the methods and compositions of the present invention are useful for both treating and preventing abnormal bone reso ⁇ tion and conditions associated therewith.
- Conditions associated with abnormal bone reso ⁇ tion include both generalized and localized bone loss.
- the creation of bone having an abnormal structure, as in Paget's disease can be associated with abnormal bone reso ⁇ tion.
- generalized bone loss means bone loss at multiple skeletal sites or throughout the skeletal system.
- localized bone loss means bone loss at one or more specific, defined skeletal sites.
- Osteoporosis is most common in post-menopausal women, wherein estrogen production has been greatly diminished. However, osteoporosis can also be steroid- induced and has been observed in males due to age. Osteoporosis can be induced by disease, e.g. rheumatoid arthritis, it can be induced by secondary causes, e.g., glucocorticoid therapy, or it can come about with no identifiable cause, i.e. idiopathic osteoporosis. In the present invention, preferred methods include the treatment or prevention of abnormal bone reso ⁇ tion in osteoporotic humans.
- Localized bone loss has been associated with periodontal disease, with bone fractures, and with periprosthetic osteolysis (in other words where bone reso ⁇ tion has occured in proximity to a prosthetic implant).
- Generalized or localized bone loss can occur from disuse, which is often a problem for those confined to a bed or a wheelchair, or for those who have an immobilized limb set in a cast or in traction.
- osteoporosis which can include post-menopausal osteoporosis, steroid-induced osteoporosis, male osteoporosis, disease-induced osteoporosis, idiopathic osteoporosis; Paget's disease; abnormally increased bone turnover; osteomalacia; periodontal disease; localized bone loss associated with periprosthetic osteolysis; and bone fractures.
- compositions and methods of the present invention are administered and caried out until the desired therapeutic effect is achieved.
- the nitrogen-containing bisphosphonate and the caspase inhibitor are generally administered concurrently.
- the nitrogen-containing bisphosphonate and the caspase inhibitor can be adminsitered sequentially.
- the caspase inhibitor is administered first.
- Murine co-cultures of osteoblasts and marrow cells are prepared using the methods of Wesolowski, et al., Exp Cell Res, (1995), 219, pp. 679-686, which is inco ⁇ orated by reference herein in its entirety.
- Bone marrow cells are harvested from 6-week-old male Balb/C mice by flushing marrow spaces of freshly isolated long bones (tibiae and femora) with ⁇ -MEM (minimal essential media) containing penicillin/streptomycin (100 I.U./ml of each and 20 mM Hepes buffer). The bone marrow cells are suspended in ⁇ -MEM and the cells are filtered through an approximately 70 ⁇ m cell strainer.
- the filtrate is centrifuged at about 300 x g for about 7 minutes.
- the resulting pellet is resuspended in ⁇ -MEM supplemented with fetal calf serum (10 % v/v) and 10 nM 1, 25-(OH)2 vitamin D3.
- fetal calf serum 10 % v/v
- These bone marrow isolates are added to sub-confluent monolayers of osteoblastic MB 1.8 cells in cell culture plates and cultured for 7 days at 37°C in the presence of 5% CO2. Culture media is replenished ever other day. Fusion of the osteoclast precursor cells from bone marrow (with each other) to form multinucleated osteoclast-like cells typically occurs after about 7 days.
- Osteoclast-like cells are enriched by sequential treatment with collagenase (1 mg/mL in phosphate buffered saline) for one hour at 37°C and EDTA (0.2 g/L in phosphate buffered saline) for 20 min at 37°C. Non-adherent cells are rinsed away by washing with phosphate buffered saline. Osteoclast-like cells which are resistant to the sequential treatments are present at about 95 % purity and are maintained in ⁇ -MEM supplemented with fetal calf serum (10 % v/v), 10 nM 1,25- (OH)2 vitamin D3, macrophage-colony-stimulating factor (5 ng/mL).
- the compounds to be evaluated are prepared as a solution of the desired concentration in ⁇ -MEM.
- examples of compounds that can be evaluated include nitrogen-containing bisphosphonates such as alendronate monosodium trihydrate, as well as compounds that block the effects of these inhibitors, such as caspase inhibitors such as Z-Val-Ala-Asp-FMK, Z-Asp-Glu-Val-Asp-FMK , and Z- Tyr-Val- Ala-Asp-FMK. Combinations of compounds can also be evaluated.
- the solutions of the compounds to be evaluated are added to the cultures for a time period of 17-24 hours. No treatment controls (controls not treated with comcpounds) are prepared by adding equivalent volumes of ⁇ -MEM to the control dishes.
- HEPES N-(2- hydroxyethyl)piperazine-N'-(2-ethansulfonic acid) or Tris buffer containing the following: ⁇ -glycerophosphate (50 mM); Na3VO4 (ImM); NaF (ImM); Microcystin LR (1 ⁇ M); leupeptin (10 ⁇ g/ml); aprotmin (10 ⁇ g/ml); phenylmethyl sulfonylfluoride (1 mM).
- HEPES N-(2- hydroxyethyl)piperazine-N'-(2-ethansulfonic acid
- Protein concentrations are determined for each lysate and 5-20 ⁇ g are loaded into each lane of a SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) gel containing Myelin Basic Protein, or another kinase substrate, which has been polymerized into the gel at a concentration between 50-400 ⁇ g/ml.
- SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- Myelin Basic Protein or another kinase substrate, which has been polymerized into the gel at a concentration between 50-400 ⁇ g/ml.
- Molecular weight standards are also loaded into one or more lanes of the gels.
- In-gel kinase assays are run according to a standard procedure based on Kameshita and Fujisawa, 1989 (Anal. Biochem. 183:139-143) and of Gotoh et al., 1990 (Eur
- Kinase reactions are then run by first soaking the gels in 20 mM HEPES, pH 7.6; 20 mM MgCl2; 2 mM DTT and then in the same buffer containing 0.02 M ATP (non-radioactive) with ca. 1000 cpm pmol 32p. ⁇ _ATP.
- the gels are then washed six times with 5% trichloroacetic acid and 1% pyrophosphate.
- the gels are then stained with Coomassie brilliant blue dye (0.125%) in 50% methanol, 10% acetic acid; destained with 30% methanol, 10% acetic acid; soaked in 2% glycerol; and dried using a gel dryer.
- the gels are then exposed to autoradiography film for times ranging from several hours to weeks.
- the bands observed in the autoradiographs representing the gels reflect kinase activities.
- Mst 1 (apparent molecular weight about 59 kDa)
- Mst 2 (apparent molecular weight about 60 kDa)
- a 34 kDa Mst kinase fragment are observed and identified by their migration as compared to the migration of molecular weight standards.
- the band intensities on the autoradiography film are quantitated by densitometry and comparisons between bands from untreated controls and bands from echistatin-treated cells provide the basis for the analyses.
- Tablets are prepared using standard mixing and formation techniques as described in U.S. Patent No. 5,358,941, to Bechard et al., issued October 25, 1994, which is inco ⁇ orated by reference herein in its entirety.
- Tablets containing about 10 mg of alendronate monosodium trihydrate, on an alendronic acid active basis, and about 0.75 mg of Z-VAD-FMK, are prepared using the following relative weights of ingredients.
- the resulting tablets are useful for administration in accordance with the methods of the present invention for treating or preventing bone reso ⁇ tion.
- tablets comprising other relative weights of alendronate, on an alendronic acid active weight basis are prepared.
- tablets containing other nitrogen-containing bisphosphonates, or mixtures thereof, at appropriate active levels are similarly prepared: e.g., cimadronate, ibandronate, risedronate, piridronate, pamidronate, zolendronate, and pharmaceutically acceptable salts thereof.
- tablets containing combinations of bisphosphonates are similarly prepared.
- tablets comprising other caspase inhibitors such as Z-Asp-
- Glu-Val- Asp-FMK or Z-Tyr-Val- Ala-Asp-FMK are prepared.
- the tablets described herein are useful for inhibiting bone reso ⁇ tion while mitigating potentially adverse gastrointestinal effects.
Abstract
Description
Claims
Priority Applications (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP99930491A EP1089750A4 (en) | 1998-06-24 | 1999-06-21 | Compositions and methods for inhibiting bone resorption |
CA002335471A CA2335471A1 (en) | 1998-06-24 | 1999-06-21 | Compositions and methods for inhibiting bone resorption |
JP2000555631A JP2002518457A (en) | 1998-06-24 | 1999-06-21 | Compositions and methods for inhibiting bone resorption |
AU47024/99A AU761138B2 (en) | 1998-06-24 | 1999-06-21 | Compositions and methods for inhibiting bone resorption |
Applications Claiming Priority (8)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US9049798P | 1998-06-24 | 1998-06-24 | |
US60/090,497 | 1998-06-24 | ||
US9291798P | 1998-07-15 | 1998-07-15 | |
US60/092,917 | 1998-07-15 | ||
GB9817170.5 | 1998-08-06 | ||
GBGB9817170.5A GB9817170D0 (en) | 1998-08-06 | 1998-08-06 | Compositions and methods for inhibiting bone resorption |
GB9819126.5 | 1998-09-02 | ||
GBGB9819126.5A GB9819126D0 (en) | 1998-09-02 | 1998-09-02 | Compositions and methods for inhibiting bone resorption |
Publications (1)
Publication Number | Publication Date |
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WO1999066945A1 true WO1999066945A1 (en) | 1999-12-29 |
Family
ID=27451814
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
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PCT/US1999/013961 WO1999066945A1 (en) | 1998-06-24 | 1999-06-21 | Compositions and methods for inhibiting bone resorption |
Country Status (5)
Country | Link |
---|---|
EP (1) | EP1089750A4 (en) |
JP (1) | JP2002518457A (en) |
AU (1) | AU761138B2 (en) |
CA (1) | CA2335471A1 (en) |
WO (1) | WO1999066945A1 (en) |
Cited By (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2000023421A1 (en) * | 1998-10-22 | 2000-04-27 | Idun Pharmaceuticals, Inc. | (SUBSTITUTED)ACYL DIPEPTIDYL INHIBITORS OF THE ICE/ced-3 FAMILY OF CYSTEINE PROTEASES |
US6492170B1 (en) | 2000-09-11 | 2002-12-10 | Isis Pharmaceuticals, Inc. | Antisense modulation of caspase 9 expression |
US6525024B1 (en) | 2000-04-17 | 2003-02-25 | Idun Pharmaceuticals, Inc. | Inhibitors of the ICE/ced-3 family of cysteine proteases |
US6566135B1 (en) | 2000-10-04 | 2003-05-20 | Isis Pharmaceuticals, Inc. | Antisense modulation of caspase 6 expression |
US7157430B2 (en) | 1998-10-22 | 2007-01-02 | Idun Pharmaceuticals, Inc. | (Substituted)acyl dipeptidyl inhibitors of the ICE/CED-3 family of cysteine proteases |
CZ303400B6 (en) * | 2000-02-01 | 2012-08-29 | Warner Chilcott Company, Llc | Selective crystallization of 3-pyridy1-1-hydroxyethylidene-1,1-bisphosphonic acid sodium as the hemipentahydrate or monohydrate and pharmaceutical composition |
-
1999
- 1999-06-21 AU AU47024/99A patent/AU761138B2/en not_active Ceased
- 1999-06-21 EP EP99930491A patent/EP1089750A4/en not_active Withdrawn
- 1999-06-21 WO PCT/US1999/013961 patent/WO1999066945A1/en not_active Application Discontinuation
- 1999-06-21 CA CA002335471A patent/CA2335471A1/en not_active Abandoned
- 1999-06-21 JP JP2000555631A patent/JP2002518457A/en not_active Withdrawn
Non-Patent Citations (6)
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COXON F P, ET AL.: "PROTEIN SYNTHESIS IS REQUIRED FOR CASPASE ACTIVATION AND INDUCTION OF APOPTOSIS BY BISPHOSPHONATE DRUGS", MOLECULAR PHARMACOLOGY, AMERICAN SOCIETY FOR PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, US, vol. 54, 1 October 1998 (1998-10-01), US, pages 631 - 638, XP002924478, ISSN: 0026-895X * |
HUGHES D E, ET AL.: "BISPHOSPHONATES PROMOTE APOPTOSIS IN MURINE OSTEOCLASTS IN VITRO AND IN VIVO", JOURNAL OF BONE AND MINERAL RESEARCH, BLACKWELL SCIENCE, INC., US, vol. 10, no. 10, 1 January 1995 (1995-01-01), US, pages 1478 - 1487, XP002924482, ISSN: 0884-0431 * |
NICHOLSON D W, THORNBERRY N A: "CASPASES: KILLER PROTEASES", TRENDS IN BIOCHEMICAL SCIENCES., ELSEVIER, HAYWARDS., GB, vol. 22, 1 August 1997 (1997-08-01), GB, pages 299 - 306, XP002924480, ISSN: 0968-0004, DOI: 10.1016/S0968-0004(97)01085-2 * |
SANDERS J L, ET AL.: "ALENDRONATE/INTERLEUKIN-1BETA COTREATMENT INCREASES INTERLEUKIN-6 IN BONE AND UMR-106 CELLS: DOSE DEPENDENCE AND RELATIONSHIP TO THE ANTIRESORPTIVE EFFECT OF ALENDRONATE", JOURNAL OF BONE AND MINERAL RESEARCH, BLACKWELL SCIENCE, INC., US, vol. 13, no. 05, 1 January 1998 (1998-01-01), US, pages 786 - 792, XP002924481, ISSN: 0884-0431, DOI: 10.1359/jbmr.1998.13.5.786 * |
See also references of EP1089750A4 * |
TAN P L J, ET AL.: "AMINOBISPHOSPHONATE INHIBITION OF INTERLEUKIN-1-INDUCED BONE RESORPTION IN MOUSE CALVARIAE", ARTHRITIS & RHEUMATISM, WILEY, US, vol. 31, no. 06, 1 June 1988 (1988-06-01), US, pages 762 - 768, XP002924479, ISSN: 0004-3591 * |
Cited By (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2000023421A1 (en) * | 1998-10-22 | 2000-04-27 | Idun Pharmaceuticals, Inc. | (SUBSTITUTED)ACYL DIPEPTIDYL INHIBITORS OF THE ICE/ced-3 FAMILY OF CYSTEINE PROTEASES |
US6242422B1 (en) | 1998-10-22 | 2001-06-05 | Idun Pharmacueticals, Inc. | (Substituted)Acyl dipeptidyl inhibitors of the ice/ced-3 family of cysteine proteases |
US7157430B2 (en) | 1998-10-22 | 2007-01-02 | Idun Pharmaceuticals, Inc. | (Substituted)acyl dipeptidyl inhibitors of the ICE/CED-3 family of cysteine proteases |
CZ303400B6 (en) * | 2000-02-01 | 2012-08-29 | Warner Chilcott Company, Llc | Selective crystallization of 3-pyridy1-1-hydroxyethylidene-1,1-bisphosphonic acid sodium as the hemipentahydrate or monohydrate and pharmaceutical composition |
US6525024B1 (en) | 2000-04-17 | 2003-02-25 | Idun Pharmaceuticals, Inc. | Inhibitors of the ICE/ced-3 family of cysteine proteases |
US6969703B2 (en) | 2000-04-17 | 2005-11-29 | Idun Pharmaceuticals, Inc. | Inhibitors of the ICE/ced-3 family of cysteine proteases |
US6492170B1 (en) | 2000-09-11 | 2002-12-10 | Isis Pharmaceuticals, Inc. | Antisense modulation of caspase 9 expression |
US6566135B1 (en) | 2000-10-04 | 2003-05-20 | Isis Pharmaceuticals, Inc. | Antisense modulation of caspase 6 expression |
Also Published As
Publication number | Publication date |
---|---|
AU4702499A (en) | 2000-01-10 |
AU761138B2 (en) | 2003-05-29 |
CA2335471A1 (en) | 1999-12-29 |
EP1089750A4 (en) | 2001-10-04 |
JP2002518457A (en) | 2002-06-25 |
EP1089750A1 (en) | 2001-04-11 |
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