OA11505A - Therapeutic combinations of (selective) estrogen receptor modulators (SERM) and growth hormone secretagogues (GHS) for treating musculoskeletal frailty. - Google Patents

Abstract

This invention is directed to pharmaceutical combination compositions and methods containing (-)-cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol or a pharmaceutically acceptable salt thereof and 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide or a pharmaceutically acceptable salt thereof, methods of using such compositions and kits containing such compositions. The compositions are useful for treating musculoskeletal frailty, including osteoporosis, osteoporotic fracture, low bone mass, frailty and low muscle mass.

Description

Ο Π 505

THERAPEUTIC COMBINATIONS OF (SELECTIVE) ESTROGEN RECEPTOR MODULATORS (SERM) AND GROWTHHORMONE SECRETAGOGUES (GHS) FOR TTREATING MUSCULOSKELETAL FRAILTY

BACKGROUND OF THE INVENTION

This invention relates to a pharmaceutical combination of a sélective5 estrogen receptor modulator (SERM) and a growth hormone secretagogue (GHS)that stimulâtes bone formation, increases bone mass, decreases sérum lipid levelsand increases muscle mass. The invention also relates to kits containing suchcombinations and the use of such combinations to treat musculoskeleta! frailty,including osteoporosis, osteoporotic fracture, low bone mass, frailty, low muscle 10 mass and the like in mammals, including humans. In particular, this inventionrelates to a combination of (-)-cis-6-phenyl-5-(4-(2’pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol or a pharmaceutically acceptable sait thereofand 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl}-1 (R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide or a 15 pharmaceutically acceptable sait thereof, kits containing such a combination andthe use of such a combination to treat musculoskeletal frailty, includingosteoporosis, osteoporotic fracture, low bone mass, frailty, low muscle mass andthe like in mammals, including humans.

Osteoporosis is a systemic skeletal disease, characterized by low bone 20 mass and détérioration of bone tissue, with a conséquent increase in bone fragilityand susceptibility to fracture. In the U.S., the condition affects more than 25million people and causes more than 1.3 million fractures each year, including500,000 spine, 250,000 hip and 240,000 wrist fractures annually. Hip fracturesare the most serious, with 5-20% of patients dying within one year, and over 50% 25 of survivors being incapacitated.

The elderly are at greatest risk of osteoporosis, and the problem is therefore predicted to increase significantly with the aging of the population.Worldwide fracture incidence is forecast to increase three-fold over the next 60years, and one study estimâtes that there will be 4.5 million hip fractures 30 worldwide in 2050.

Although both men and women are susceptible to musculoskeletal frailty, including osteoporosis, women are at greater risk of osteoporosis than men.

Women expérience a Sharp accélération of bone loss immediately following 011505 ménopausé. Other factors that increase bone loss leading to osteoporosis indude smoking, alcohol abuse, a sedentary lifestyle and low calcium intake.

Estrogen is the agent of choice in preventing osteoporosis or post menopausal bone loss in women. In addition, Black, et al. in EP 0605193A1 5 report that estrogen, particularly when taken orally, lowers plasma levels of LDLand raises those of the bénéficiai high density lipoproteins (HDL’s). Long-termestrogen therapy, however, has been implicated in a variety of disorders, indudingan increase in the risk of uterine cancer, endométrial cancer and possibly breastcancer, causing many women to either avoid this treatment or take the médication 10 for only a short period of time. Although the risk of endométrial cancer is thoughtto be reduced by a concurrent use of a progestérone, there is still concem aboutpossible increased risk of breast cancer with the use of estrogen. Recentlysuggested therapeutic regimens, which seek to lessen the cancer risk, such asadministering combinations of progestérone and estrogen, cause the patient to 15 expérience unacceptable bleeding. Furthermore, combining progestérone withestrogen seems to blunt the sérum cholestérol lowering effects of estrogen. Thesignificant undesirable side effects associated with estrogen therapy support theneed to develop alternative thérapies for osteoporosis that hâve the désirablebénéficiai effect on sérum LDL but do not cause undesirable side effects. 20 Recently, a number of sélective estrogen receptor modulators hâve been proposed for treatment of osteoporosis. It has been reported (OsteoporosisConférence Scrip No. 1812/13 April 16/20, 1993, p. 29) that raloxifene, 6-hydroxy-2-(4-hydroxyphenyl)-3-[4-(2-piperidinoethoxy) benzoyl] benzo[b] thiophene, mimicsthe favorable action of estrogen on bone and lipids but, unlike estrogen, has 25 minimal uterine stimulatory effect. [Black, LJ. et al., Raloxifene (LY139481 Hcl)Prevents Bone Loss and Reduces Sérum Cholestérol Without Causing UterineHypertrophy in Ovariectomized Rats, J. Clin. Invest., 1994, 93:63-69 and Delmas,P.D. et al., Effects of Raloxifene on Bone Minerai Density, Sérum CholestérolConcentration, and Uterine Endometrium in Postmenopausal Women, New 30 England Journal of Medicine, 1997, 337:1641-1647],

Agents such as droloxifene, U.S. pat. no. 5,254,595, prevent bone loss and thereby reduce the risk of fracture without estrogen's side effects. However, estrogen and estrogen agonists alone are only expected to reduce the fracture risk 01 1 505 by about 50% leaving approximately 50% of ostéopénie women still at risk for anosteoporotic fracture.

Commonly assigned U.S. pat. no. 5,552,412, which is incorporated hereinby reference, discloses SERM compounds of the formula

wherein the variables are defined as set forth therein.

Growth hormone (GH), which is secreted from the pituitary gland,stimulâtes growth of ail tissues of the body that are capable of growing. Inaddition, GH is known to hâve the following basic effects on the metabolic processof the body: 1. Increased rate of protein synthesis in substantially ail cells of thebody; 2. Decreased rate of carbohydrate utilization in cells of the body; 3. Increased mobilization of free fatty acids and use of fatty acids forenergy.

Deficiency in GH résulte in a variety of medical disorders. In children, itcauses dwarfism. In adults, the conséquences of acquired GH deficiency includeprofound réduction in lean body mass and concomitant increase in total body fat,particulariy in the truncal région. Decreased skeletal and cardiac muscle massand muscle strength lead to a significant réduction in exercise capacity. Bonedensity is also reduced. Administration of exogenous GH has been shown toreverse many of the metabolic changes. Additional benefits of therapy hâveincluded réduction in LDL cholestérol and improved psychological well-being.

In cases where increased levels of GH were desired, the problem wasgenerally solved by providing exogenous GH or- by administering an agent whichstimulated GH production and/or release. In either case the peptidyl nature of thecompound necessitated that it be administered by injection. Initially the source of 011505 GH was the extraction of the pituitary glands of cadavers. This resulted in an expensive product, and carried with it the risk that a disease associated with the source of the pituitary gland could be transmitted to the récipient of the GH (e.g.,

Jacob-Creutzfeld disease). Recently, recombinant GH has become available 5 which, while no longer carrying any risk of disease transmission, is still a veryexpensive product which must be given by injection or by a nasal spray.

Most GH deficiencies are caused by defects in GH release, not primarydefects in pituitary synthesis of GH. Therefore, an alternative strategy fornormalizing sérum GH levels is by stimulating its release from somatotrophs. 10 Increasing GH sécrétion can be achieved by stimulating or inhibiting variousneurotransmitter Systems in the brain and hypothalamus. As a resuit, thedevelopment of synthetic GH-releasing agents to stimuiate pituitary GH sécrétionare being pursued, and may hâve several advantages over expensive andinconvénient GH replacement therapy. By acting along physiologie regulatory 15 pathways, the most désirable agents would stimuiate pulsatile GH sécrétion, andexcessive levels of GH that hâve been associated with the undesirable sideeffects of exogenous GH administration would be avoided by virtue of intactnégative feedback loops.

Physiologie and pharmacologie stimulators of GH sécrétion include 20 arginine, L-3,4-dihydroxyphenylalanine (L-DOPA), glucagon, vasopressin, andinsulin induced hypoglycemia, as well as activities such as sleep and exercise,indirectly cause GH to be released from the pituitary by acting in some fashion onthe hypothalamus perhaps either to decrease somatostatin sécrétion or toincrease the sécrétion of the known secretagogue GH releasing factor (GHRF) or 25 an unknown endogenous GH-releasing hormone or ail of these.

Commonly assigned International Patent Application Publication Number WO97/24369, designating, inter alia, the United States, discloses GHsecretagogues of the formula 01 1 505

wherein the variables are defined as set forth therein. International PatentApplication Number WO97/24369 is incorporated herein by reference.

Tang et al., Restoring and Maintaininq Bone in Ostéogénie Female Rat 5 Skeleton; I. Changes in Bone Mass and Structure, J. Bone Minerai Research 7 (9),p1093-1104, 1992 discloses data for the lose, restore and maintain (LRM)concept, a practical approach for reversing existing osteoporosis. The LRMconcept uses anabolic agents to restore bone mass and architecture (+ phase)and then switches to an agent with the estabiished ability to maintain bone mass, 10 to keep the new bone (+/- phase). The rat study utilized PGE2 and risedronate, abisphosphonate, to show that most of the new cancellous and cortical boneinduced by PGE2 can be maintained for at least 60 days after discontinuing PGE2by administering risedronate.

Shen et al., Effects of Reciprocal Treatment with Estrogen and Estrogen 15 plus Parathyroid Hormone on Bone Structure and Strength in OvariectomizedRats, J. Clinical Investigation, 1995, 96:2331-2338 discloses data for thecombination and/or sequential use of anti-resorptive agents and anabolic agentsfor the treatment of osteoporosis.

Commonly assigned International Patent Application Publication Number 20 WO97/31640, designating, inter alia, the United States, discloses the use of certain GH secretagogues in combination with certain SERMS to treatosteoporosis. International Patent Application Publication Number WO97/31640 isincorporated herein by. reference.

SUMMARY OF THE INVENTION 25 This invention is directed to a pharmaceutical composition comprising: a. a first compound, said first compound being (-j-cis-ô-phenyl-S^A-^- pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol or a pharmaceutically acceptable sait thereof; and b 01 1 505 b. s second compound, said second compound being 2-amino-N-{2- (3a(R)-benzyl-2-methyl-3-cxo-2,3, 3a, 4.6,7-hexahydrc-pyrazcio-(4,3-c]pyridin-5- yl)-1(R)-benzy!oxymethyl-2-oxo-ethyl)-iscbutyramide or a pharmaceutically sccaptable sait thereof.

This invention is furiher cirected to a pharmaceuticai composition as recitedin the immecistely preceding paragraph additicnally comprising a pharmaceuticai carrier.

This invention is sise directed to a pharmaceuticai composition ssdescribed in either cf the first Mo paraçrsphs of this summary wherein said first 10 compound is (-)-cis-6-phenyi-5-(4-(2-pyrrolid:n-'i-yi-ethcxy}-phenyl)-5,6,7,5- tetîahydronaphihaiene-2-oi D-tsrtfête and saie second compound is 2-amino-N-(2-(3a(R)-benzy!-2-meLhy!-3-oxo-2,3, 3a, 4,6,7-hexshyaro-pyrazcio-{4,2-c}pyridin-5-yl)-1(R}-benzyioxymethyl-2-oxo-ethy!)-isobutyremide L-tartrste.

This invention is still furiher directed to a method, cesionated Method A, for15 treating a mammal suffering from muscuioskeietai fraiity comprising administerinçto said mammal a pharmaceuticai composition as recited in any of the first three paragraphs of this summary. A preferred method within Method A, designated Method B, is wherein themammal is suffering from osteoporosis. 20 Another preferred methoc within Method A, designated Method C, is wherein mammal is suffering from osteotomy, chiidhcod idiopathic bone loss crbone loss associated with periodontitis.

This invention is still furiher directed to a method, designated Method A1,for treating a mammal suffering from muscuioskeietai fraiity comprising 25 administering to said mammal a. a first compound, said first compound being (-)-cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6l7,8-tetrahydronaphthaiene-2-ol or apharmaceutically acceptable sait thereof; and b. a second compound, said second compound being 2-amino-N-(2-30 Qa(R)-benzyl-2-methyl-3-oxo-2;3,3a,4,6,7-hexahydro-pyrazolo[4,3-c]pyridin-5- yl)-l(R)-benzyloxymethyl)-2-oxo-ethyl)-isobutyramide or a· pharmaceuticallyacceptable sait thereof. 01 1 50 5 10 15 20 25 30

This invention is parfculariy directed to a method of Method A’ wherein thefirst compound and the second compcund are administered substantiallysimultanecusiy.

This invention is sise particulariy directed to s methcd ci Methcd A1,hereinafter terrned Method D, wherein the second compound is administered fer s peried cf front about three menths to about three years.

This invention is more psrticulariy directed to a methcd c-f Method D foüowed fcv administration of the first compound for a peried of from about three menths te about three years without the administration cf the second compcunddurino the pericc cf from about three menths to about three years.

This invention is also more particulariy directed to a methcd cf Method Dfoiiowed bv administration of the first compcund for a period oreater than aboutthree years without the administration of the second compound durino the oreaterthan atout three year perioc.

This invention is also directed to a method, hereinafter terrned Method E,for treating muscuioske'.eta! fraiity in a mammal suffering therefrom comprisincadministering to said msmmsi a therapeutically effective ameunt of a compositionas recited in sny of the first three paraaraphs of this summary. A preferred method within Method E is wherein bone healing foîlowingfacial reconstruction, maxiilary reconstruction or mandibuiar reconstruction isenhanced, vertébral synostosis is induced, long bone extension is enhanced, thehealing rate of a bone graft or a long bone fracture is enhanced or prostheticinorewth is enhanced. Additionally preferred is a method comprising administeringto said mammal a. a first compound, said first compound being (-)-cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyf}-5,6,7,&-tetrahydronaphthalene-2-ol or apharmaceutically acceptable sait thereof; and b. a second compound, said second compound being 2-amino-N-(2-.(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo[4,3-c]pyridin-5-yl)-l(R)-benzyloxymethyl)~2-oxo-ethyl)-isobutyramide or a pharmaceuticallyacceptable sait thereof.

This invention is also directed to a methcd for increasing muscle mass in a mammal comprisinc administering to said mammal a muscle mass increasing 0115 0 5 effective amount or a composition as recited in any of the first three paragraphe ofthis summary. Additionally preferred is a method comprising administerina to saidmammal a. a first compound, said first compcunc being (-)-cis-6-phenyl-5-(4-{2- 5 pyrrolid;n-1-yl-ethcxy)-phenyl)-5,6,7,8-tetrahydronaphthsiene-2-cl or a pharmaceuticaily acceptable sait thereof; and b. a second compound, said second compound being 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4J6,7-hexahydro-pyrazolo[4,3-c]pyridin-5-'yl)-l(R)-benzyloxymethyl)-2-oxo-eihy!)-isobutyramide or a pharmaceuticaily 10 | acceptable sait thereof.

In ail of the methods of this invention, it is particulariy preferred triât themammal is a human.

This invention is sise directed te a kit comprising a Veatment for a mammalsuffering frem musculoskeletal frailty comprisina: 15 a. s therapeuticallv effective amount ci (-)-cis-€-phenyl-5-(4-(2-pyrroÎidin-1- yi-ethcxy)-phenyl)-5,6,7,8-îe!rahyGronaphihalene-2-cl or a pharmaceuticailyacceptable sait thereof and a pharmaceuticaily acceptable carrier in a first unitdosage form; b. s therspeuticslly effective amount of 2-sminc-N-(2-(3a(R)-benzyl-2- 20 methyl-3-oxo-2,3, 3a, 4,6,7-hexshydro-pyrazolo-{4,3-c]pyridin-5-yl)-1(R)- benzyloxymethyl-2-Gxo-ethy!)-iscbutyramide or a pharmaceuticaily acceptable saitthereof and a pharmaceuticaily acceptable carrier in a second unit dosage form;and c. a container. 25 This invention is particulariy directed to a kit as described in the immediately preceding paragraph wherein said first unit dosage form comprises (->cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8- tetrahydronaphthalene-2-ol D-tartrate and said second unit dosage form comprises2-amino-N-(2-(3a(R)-benzyt'2-methyl-3-oxô-2,3, 3s, 4,6,7-hexahydro-pyrazolo- 30 [4,3-c]pyridin-5-yl)-1 (R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide L-tartrate.

In ail of the compositions, methods and kits of this invention, it is particulariy preferred that the D-tartrate sait of (-)-cis-6-phenyi-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-cl is used and that the L- 011 505 tartrate sait of 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7- hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)- isobutyramide is used.

The phrase "condition which présents with low bone mass' refers to a 5 condition where the level of bone mass is below the âge spécifie normal asdefined in standards by the World Health Organization "Assessment of FractureRisk and its Application to Screening for Postmenopausal Osteoporosis (1994),Report of a World Health Organization Study Group. World Health OrganizationTechnical Sériés 843”. Çhildhood idiopathic and primary osteoporosis are also 10 included. Included in the treatment of osteoporosis is the prévention oratténuation of long term complications such as curvature of the spine, loss ofheight, prosthetic surgery, and prévention of prostate malfunctioning. Alsoincluded is increasing the bone fracture healing rate and enhancing the rate ofsuccessful bone grafts. Also included is periodontal disease and alveolar bone 15 loss.

The phrase "condition which présents with low bone mass" also refers to arnammal known to hâve a significantly higher than average chance of developingsuch diseases as are described above including osteoporosis (e.g., post-menopausal women, men over the âge of 60, and persons being treated with 20 drugs known to cause osteoporosis as a side effect (such as glucocorticoid)).

Those skilled in the art will recognize that the term bone mass actuallyrefers to bone mass per unit area which is sometimes (although not strictlycorrectly) referred to as bone minerai density.

The phrase “musculoskeletal frailty" refers to a condition wherein a subject25 has low bone mass and/or low muscle mass, and includes such diseases,disorders and conditions such as, but not limrted to, conditions which présent withlow bone mass, osteoporosis, conditions which présent with low muscle mass,osteotomy, çhildhood idiopathic bone loss, bone loss associated with periodontitis,bone healing following facial reconstruction, maxillary reconstruction, mandibular 30 reconstruction and bone fracture. Further, musculoskeletal frailty encompasses such conditions as interfaces between newly attached prostheses and bone which require bone ingrowth. 10 011505

The term "treating", "treat" or "treatment" as used herein includes curative,preventative (e.g., prophylactic) and palliative treatment.

The parenthetical négative or positive sign used herein in the nomenclaturedénotés the direction plane polarized light is rotated by the particular stereoisomer.

The compositions of this invention may include hydrates of the compoundsused therein.

The phamnaceutical compositions and methods of this invention resuit in amore rapid and higher magnitude bone mass gain than is achievable with thesame doses of (-)-c/'s-6-phenyl-5-[4-(2-pyrrolidin-1-yl-ethoxy)-phenyl]-5,6,7,8-tetrahydro-naphthalene-2-ol as described above alone or 2-amino-N-[2-(3a-(R)-benzyl-2-methyl-3Oxo-2I3,3a,4l6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5yl)-1-(R)-benzyloxymethyl-2-oxo-ethyl]-isobutyramide as described above alone. Further,these combinations increase bone density and muscle mass while at the sametime redudng fat mass and total sérum cholestérol. Thus, these combinationsincrease bone mass and decrease fracture rates to a greater extent than isachievable through use of either agent alone. This invention makes a significantcontribution to the art by providing compositions and methods that increase andmaintain bone mass resulting in prévention, retardation, and/or régression ofosteoporosis and related bone disorders.

Other features and advantages will be apparent from the spécification anddaims which describe the invention.

DETAILED DESCRIPTION OF THE INVENTION

The First compound of this invention is (-)-c/s-8-phenyl-5-[4-(2-pymolidin-1-yl-ethoxy)-phenyl]-5,6,7,8-tetrahydro-naphthalene-2-ol or a pharmaceuticallyacceptable sait thereof, which has the structure of Formula I: 11 01 1 505

(-)-C/s-6-phenyl-5-[4-(2-pyrrolidin-1-yl-ethoxy)-phenyl]-5,6,7,8-tetrahydro-naphthalene-2-ol and the pharmaceutically acceptable salts thereof are prepared 5 as described in commonly assigned US Patent Number 5,552,412, which isreferenced above. (-)-C/s-6-phenyl·5-[4-(2-pyrrolidin-1-yl-ethoxy)-pheny^}-5I6,7,8-tetrahydro-naphthalene-2-ol D-tartrate is prepared as set forth in the immediately preeedingparagraph or, altematively, as set forth in International Patent Application 10 Publication Number WO97/16434, designating the United States and which isincorporated herein by reference.

The second compound of this invention is 2-amino-N-[2-(3a-(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1-(R)-benzyloxymethyl-2-oxo-ethyl]-isobutyramide or a pharmaceutically acceptable sait 15 thereof, which has the structure of Formula II:

II 12 01 1 505 2-Amino-N-[2-(3a-(R)-benzyl-2-methyl-3-oxo-2,3.3a, 4,6,7-hexahydro-pyrazolo-[4,3-c]pyridiri’5-yl)-1 -(R)-benzyloxymethyl-2-oxo-ethyl]-isobutyramide andpharmaceutically acceptable salts thereof are prepared as set forth in commonlyassigned Internationa! Patent Application Publication Number WO97/24369, which 5 is referenced above.

In addition, when the compounds or the pharmaceutically acceptable saltsthereof of this invention form hydrates or solvatés they are also within the scope ofthe invention.

The pharmaceutical combinations and methods of this invention are ail10 adapted to therapeutic use as agents that either activate bone turnover or prevent bone résorption or increase bone formation in mammals, particularly humans.Since these functions are closely related to the development of osteoporosis andbone related disorders, these combinations, by virtue of their action on bone,prevent, arrest, regress or reverse osteoporosis. 15 The utility of the compositions and methods of the présent invention as medical agents in the treatment of musculoskeletal frailty (e.g., conditions whichprésent with low bone mass or low muscle mass including osteoporosis) inmammals (e.g. humans) is demonstrated by the activity of the compounds of thisinvention in conventional assays as set forth in U.S. Patent Number 5,552,412 and 20 International Patent Application Publication Number WO97/2436Ô. Furtherevidence of the utility of the instant combination is set forth in Example One below.Such assays also provide a means whereby the activities of the compounds ofthis invention can be compared between themselves and with the activities ofother known compounds. The résulte of these compensons are useful for 25 determining dosage levels in mammals, including humans, for the treatment ofsuch diseases.

Administration of the compounds of this invention can be via any methodwhich delivers a compound of the combination of this invention systemically and/orlocally. These methods include oral routes, parentéral, intraduodenal routes, etc. 30 Generally, the compounds of this invention are administered orally, but parentéral administration (e.g., intravenous, intramuscular, transcutaneous, subcutaneaus or intramedullary) may be utilized, for example, where oral administration is inappropriate for the instant target or where the patient is unable to ingest the 13 011505 drug. The two different compounds of this invention can be co-administeredsimultaneously or sequentially in any order, or a single pharmaceuticalcomposition comprising a first compound as described above and a secondcompound as described above in a pharmaceutically acceptable carrier can be 5 administered.

In any event the amount and timing of compounds administered will, ofcourse, be dépendent on the subject being treated, on the severity of the affliction,on the manner of administration and on the judgment of the prescribing physician.Thus, because of patient to patient variability, the dosages given below are a 10 guideline and the physician may titrate doses of the drug to achieve the activity(e.g., bone mass augmentation) that the physician considers appropriate for theindividual patient. In considering the degree of activity desired, the physician mustbalance a variety of factors such as bone mass starting level, âge of the patient,presence of preexisting disease, as well as presence of other diseases (e.g., 15 cardiovascular). For example, the administration of (-)-cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol can providecardiovascular beneflts, particularly for post-menopausal women. The followingparagraphs provide preferred dosage ranges for the various components of thisinvention. 20 An effective dosage for (-)-c/s-6-phenyl-5-[4-(2-pyrrolidin-1-yl-ethoxy)- phenyl]-5,6,7,8-tetrahydro-naphthalene-2-ol is in the range of 0.0001 to 100mg/kg/day, preferably 0.001 to 10 mg/kg/day.

An effective dosage for 2-amino-N-(2-(3a(R)-benzyi-2-methyl-3-oxo-2,3, 3a,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)- 25 isobutyramide is in the range of 0.0001 to 100 mg/kg/day, preferably 0.01 to 5mg/kg/day.

Where the tartrate sait or other pharmaceutically acceptable sait of eitherof the above compounds is used in this invention, the skilled person will be able tocalculate effective dosage amounts by calculating the molecular weight of the sait 30 form and performing simple stoichiometric ratios.

The-compounds of the présent invention are generally administered in theform of a pharmaceutical composition comprising at least one of the compounds orpharmaceutically acceptable salts thereof of this invention together with a 14 011505 pharmaceutically acceptable vehicle or diluent. Thus, the compounds andpharmaceutically acceptable salts thereof of this invention can be administeredseparately or together in any conventional oral, parentéral or transdermal dosageform. When administered separately, the administration of the other compound or 5 a pharmaceutically acceptable sait thereof of the invention follows.

For oral administration a pharmaceutical composition can take the form of solutions, suspensions, tablets, pills, capsules, powders, and the like. Tabletscontaining various excipients such as sodium citrate, calcium carbonate andcalcium phosphate are employed along with various disintegrants such as starch 10 and preferably potato or tapioca starch and certain complex silicates, together withbinding agents such as polyvinylpyrrolidone, sucrose, gelatin and acacia.Additionally, lubricating agents such as magnésium stéarate, sodium lauryl sulfateand talc are often useful for tabletting purposes. Solid compositions of a similartype are also employed as fillers in soft and hard-filled gelatin capsules; preferred 15 materials in this connection also include lactose or milk sugar as well as highmolecular weight polyethylene glycols. When aqueous suspensions and/or élixirsare desired for oral administration, the compounds or pharmaceutically aceptablesalts thereof of this invention can be combined with various sweetening agents,flavoring agents, coloring agents, emulsifying agents and/or suspending agents, 20 as well as such diluents as water, éthanol, propylene glycol, glycerin and variouslike combinations thereof.

For purposes of parentéral administration, solutions in sesame or peanut oilor in aqueous propylene glycol can be employed, as well as stérile aqueoussolutions of the corresponding water-soluble salts. Such aqueous solutions may 25 be suitably buffered, if necessary, and the liquid diluent first rendered isotonie withsufficient saline or glucose. These aqueous solutions are especially suitable forintravenous, intramuscular, subeutaneous and intraperitoneal injection purposes.In this connection, the stérile aqueous media employed are ail readity obtainableby standard techniques well-known to those skilled in the art 30 For purposes of transdermal (e.g.,topical) administration, dilute stérile, aqueous or partially aqueous solutions (usually in about 0.1% to 5% concentration), otherwise similar to the above parentéral solutions, are prepared. 15 01 1 505

Methods of preparing various pharmaceutical compositions with a certain amount of each active ingrédient are known, or will be apparent in light of this disclosure, to those skilled in this art. For examples, see Reminqtorïs

Pharmaceutical Sciences. Mack Publishing Company, Easton, Pa., 19th Edition (1990).

Pharmaceutical compositions according to the invention may contain 0.1%-95% of a combination of the compounds or pharmaceutically acceptable saltsthereof of this invention, preferably 1%-70%. In any event, the composition orformulation to be administered will contain a quantity of the compounds orpharmaceutically acceptable salts thereof of the invention in an amount effectiveto treat the disease/condition of the subject being treated.

Since the présent invention relates to treatment with a combination of thetwo active ingrédients which may be administered separately, the invention alsorelates to combining separate pharmaceutical compositions in kit form. The kitincludes two separate pharmaceutical compositions: (-)-cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol or apharmaceutically acceptable sait thereof and 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7-hexahydro-pyrazolo-I4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide or a pharmaceutically acceptable saitthereof. The kit includes a container for containing the separate compositions suchas a divided bottle or a divided foil packet, however, the separate compositionsmay also be contained within a single, undivided container. Typically the kitincludes directions for the administration of the separate components. The kitform is particularty advantageous when the separate components are preferablyadministered in different dosage forrns (e.g., oral and parentéral), are administeredat different dosage intervals, or when titration of the individual components of thecombination is desired by the prescribing physician.

An example of such a kit is a so-called blister pack. Blister packs are wellknown in the packaging industry and are being widely used for the packaging ofpharmaceutical unit dosage forrns (tablets, capsules, and the like). Blister packsgenerally consist of a sheet of relatively stiff material covered with a foil of apreferably transparent plastic material. During the packaging process recessesare formed in the plastic foil. The recesses hâve the size and shape of the tablets 16 011 505 or capsules to be packed. Next, the tablets or capsules are placed in the recesses and the sheet of relatively stiff material is sealed against the plastic foi! at the face of the foil which is opposite from the direction in which the recesses were formed.

As a resuit, the tablets or capsules are sealed in the recesses between the plastic 5 foil and the sheet. Preferably the strength of the sheet is such that the tablets orcapsules can be removed from the blister pack by manually applying pressure onthe recesses whereby an opening is formed in the sheet at the place of the recess.The tablet or capsule can then be removed via said opening.

It is désirable to provide a memory aid on a card insert, e.g., in the form of 10 numbers next to the tablets or capsules whereby the numbers correspond with thedays of the regimen which the tablets or capsules so specified should be ingested.Another example of such a memory aid is a calendar printed on the card e.g., asfollows "First Week, Monday, Tuesday, ...etc.... Second Week, Monday,Tuesday,..." etc. Other variations of memory aids will be readily apparent A 'daily 15 dose" can be a single tablet or capsule or several pills or capsules to be taken ona given day. Also a daily dose of SERM can consist of one tablet or capsule whilea daily dose of a GH secretagogue can consist of several tablets or capsules. Thememory aid should refiect this.

In another spécifie embodiment of the invention a dispenser designed to 20 dispense the daily doses one at a time in the order of their intended use isprovided. Preferably, the dispenser is equipped with a memory-aid, so as tofurther facilitate compliance with the regimen. An example of such a memory-aidis a mechanical counter which indicates the number of daily doses that has beendispensed. Another example of such a memory-aid is a battery-powered micro- 25 chip memory coupled with a liquid crystal readout, or audible reminder signalwhich, for example, reads out the date that the last daily dose has been takenand/or reminds one when the next dose is to be taken.

The following assay is used to show that the combination and methods ofthis invention increases lean body mass and decreases fat body mass whereas 30 the GH secretagogue alone would be expected to decrease fat body mass with no change in lean body mass and the SERM alone would be expected ίο increase both lean and fat body mass. Further, the combination increases bone density and decreases total sérum cholestérol. 17 01 1 505

Example One

Female S-D rats (Karlan) were sham-operated or ovariectomized (OVX) at3.5 months of âge. Drug administration started when the rats were 9 months ofâge and 5.5 months post-surgery. The sham-operated rats received daily gavageof vehicle (10% éthanol in water), while the OVX rats received daily gavage ofvehide, or 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo-[4,3-c)pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxoethyl)-isobutyramide at 5mg/kg/d alone, or (-)cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol at 0.1 mg/kg/d alone, or co-treatment of 2-amino-N-(2-(3a(R)-benzy!-2-methyl-3-oxo-2,3,38,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1 (R)-benzyloxymethyl-2-oxoethyl)-isobutyramide and (-)cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphtha!ene-2-ol for 4 weeks. Inthe combination group, 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxoethyl)-isobutyramide was given 2 hours prior to (-)cis-ô-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol. There were 8 to 10 rats pereach subgroup. AH rats were given subcutaneous injections with 10 mg/kg ofcalcein (Sigma Chemical Co., St. Louis, MO) on 13 and 3 days before autopsy. Itwill be recognized by those skilled in the art that the compounds used in this assaymay be administered in the form of a pharmaceutically acceptable sait and that thedosage amount can be readily determined by calculating the molecular weight ofthe sait and perfomning simple ratios.

Before autopsy on the terminal day of the assay, ail rats underketamine/xylazine anesthésia underwent dual-energy X-ray absorptiometry (DXA,QDR-1000/W, Hologic Inc., Waltham, MA) equipped with Rat Whole Body Scansoftware (Hologic Inc., Waltham, MA) for lean and fat body mass détermination.The rats were then autopsied and blood was obtained by cardiac puncture. Totalsérum cholestérol was determined using a high performance cholestérolcolorometic assay (Boehringer Mannheim Biochemicals, Indianapolis, IN). Thebody weight gain was calculated as body weight at autopsy minus body weight atday 0. The uterine wet weight was determined immediately at autopsy.

The right fémur from each rat was removed at autopsy and scanned using dual energy χ-ray absorptiometry (DXA, QDR 1000/W, Hologic Inc., Waltham, MA) 18 01 1 505 equipped with "Régional High Resolution Scan” software (Holcgic Inc., Waltham,MA). The scan field size was 5.08 x 1.902 cm, resolution is 0.0254 x 0.0127 cmand scan speed was 7.25 mm/second. The fémoral scan images were analyzedand total fémoral bone area, bone minerai content, and bone minerai density were 5 determined according to the method described in H. Z. Ke et al.. Droloxifene, aNew Estrogen Antagonist/Agonist, Prevents Bone Loss in Ovariectomized Rats.ENDOCRINOLOGY 136;2435-2441, 1995.

Study Results and Discussion

Compared to the Controls, 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo- 10 2,3,3a,4,6,7-hexahydro-pyrazolo-t4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2- oxoethyl)-isobutyramide atone increased both lean and fat body mass, while (-)cis- 6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,&-tetrahydronaphthalene-2-olalone decreased fat body mass with no change in lean body mass. Combinationof 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo- 15 [4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxoethyl)-isobutyramide and (-)cis-6-phenyl’5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7l8-tetrahydronaphthalene-2-olincreased lean body mass and decreased fat body mass. Therefore, combinationof both compounds hâve a better body composition profile than 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)- 20 1(R)-benzyloxymethyl-2-oxoethyl)-isobutyramide alone or (-)ds-6-phenyl-5-(4-(2-pyrTolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol alone. A 5-6% increase in total fémoral bone area was observed in subjectsreceiving the combination relative to those subjects who received placebo. Thisresuit is similar to the increase in total fémoral bone area which was observed in 25 subjects receiving either (-)cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol or 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxoethyl)-isobutyramide and (-)ds-6-phenyl-5-(4-(2-pynOlidin-1 -yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol alone. Total fémoral bone minerai content 30 increased by 8.5% in 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3a14,6,7- hexahydro-pyrazolo-(4,3-c]pyndin-5-yl)-1(R)-benzyloxymethyl-2-oxoethyl>- isobutyramide alone and 7.7% in (-)cis-6-phenyL5-(4-(2-pynOlidin-1-yl-ethoxy)- phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol alone. However, in the combination 19 011505 group, total fémoral bone minerai content increased by 12.5%, which was asignificant increase compared to either alone. A similar pattern was found in thetotal fémoral bone minerai density.

Total sérum cholestérol decreased in (-)cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl- 5 ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol alone group and thecombination group.

These data indicated that combination of 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3,3al4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxoethyl)-isobutyramide and (-)cis-6-phenyl-5-(4-(2-pymolidin-1- 10 yl-ethoxy)-phenyl)-5,6,7,&-tetrahydronaphthalene-2-ol, hâve multiple benefits.These benefits include an increase in lean mass and a decrease in fat mass andsérum lipid. Further, an increase in bone mass was observed.

It should be understood that the invention is not limited to the particularembodiments described herein, but that various changes and modifications may 15 be made without departing from the spirit and scope of this invention as defined bythe following daims.

Claims (12)

1. A pharmaceutical composition comprising: a. a first compound, said first compound being (-)-cis-6-phenyl-5-(4-(2- pymolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol or a 5 pharmaceutically acceptable sait thereof; and b. a second compound, said second compound being 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide or a pharmaceutically acceptable saitthereof. 10

2. A pharmaceutical composition of claim 1 additionally comprising a pharmaceutical carrier.

01 1 505 20 PRODUCT ClAIMS

3. A pharmaceutical composition of claim 1 wherein said firstcompound is (-}-cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol D-tartrate and said second compound is 2-amino-N-(2- 15 (3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1 (R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide L-tartrate.

4. Use of a pharmaceutical composition of claim 1 to préparé amédicament for treating a mammal suffering from musculoskeletal frailty.

5. A use of claim 4 wherein said first compound is (-)-cis-6-phenyl-5-20 (4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol D-tartrate and said second compound is 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a,4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide L-tartrate.

6. A use of claim 4 wherein said mammal is suffering from25 osteoporosis.

7. A use of claim 4 wherein osteotomy, childhood idiopathic bone lossor bone loss associated with periodontitis is treated.

8. The use of claim 4 wherein bone healing following facialreconstruction, maxillary reconstruction or mandibular reconstruction is treated, 30 vertébral synostosis is induced or long bone extension is enhanced, the healingrate of a bone graft is enhanced or prosthetic ingrowth is enhanced,

9. The use of claim 8 wherein a bone fracture is treated in a human.

10. A use of claim 6 wherein osteoporosis is treated in a human. 21 01 1 505

11. A kit comprising: a. (-)-cis-6-phenyl-5-(4-(2-pymolidin-1-yi-ethoxy)-phenyi)-5,6,7,8- tetrahydronaphthalene-2-ol or a pharmaceutically acceptable sait îhereof and apharmaceutically acceptable carrier of diluent in a first unit dosage form; 5 b. 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7- hexahydro-pyrazolo-{4,3-c]pyridin-5-yl)-1(R)-benzyloxymethyl-2-oxo-ethyl)-isobutyramide or a pharmaceutically acceptable sait thereof and apharmaceutically acceptable carrier or diluent in a second unit dosage form; and c. a container. 10

12. A kit of claim 11 wherein said first unit dosage form comprises (-)- cis-6-phenyl-5-(4-(2-pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydronaphthalene-2-ol D-tartrate and said second unit dosage form comprises 2-amino-N-(2-(3a(R)-benzyl-2-methyl-3-oxo-2,3, 3a, 4,6,7-hexahydro-pyrazolo-[4,3-c]pyridin-5-yl)-1 (R)-benzyloxymethyl-2-oxo-ethyi)-isobutyramide L-tartrate. 15 20 25 30